Probiotic compositions containing clostridiales for inhibiting inflammation

ABSTRACT

Pharmaceutical compositions containing microbial entities are described herein. The pharmaceutical compositions may optionally contain or be used in conjunction with one or more prebiotics. Uses of the pharmaceutical compositions to treat or prevent disorders of the local or systemic microbiome in a subject are also provided.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/438,271, filed Feb. 21, 2017 which, in turn, is a continuation ofU.S. patent application Ser. No. 14/952,895, filed Nov. 25, 2015, which,in turn, claims priority to U.S. Provisional Patent Application No.62/084,536, filed Nov. 25, 2014; U.S. Provisional Patent Application No.62/084,537, filed Nov. 25, 2014; U.S. Provisional Patent Application No.62/084,540, filed Nov. 25, 2014; U.S. Provisional Patent Application No.62/117,632, filed Feb. 18, 2015; U.S. Provisional Patent Application No.62/117,637, filed Feb. 18, 2015; U.S. Provisional Patent Application No.62/117,639, filed Feb. 18, 2015; U.S. Provisional Patent Application No.62/162,562, filed May 15, 2015; and U.S. Provisional Patent ApplicationNo. 62/257,714, filed Nov. 19, 2015. The entire contents of each of theforegoing applications are incorporated herein by reference.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in ASCII format and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Dec. 13, 2018, isnamed 126383_02104_SL.txt and is 4,147,567 bytes in size.

BACKGROUND

Humans and other mammals have numerous microbial niches, andinterventions to modulate the microbiota thereof have been focused onantibiotics (which effect largely non-specific eradication of themicrobiota in an effort to target a pathogen), probiotics (largely inthe form of lactic acid-producing bacteria in food products), prebiotics(stimulatory materials, primarily carbohydrates, that increase bacterialgrowth and/or activity), and synbiotics (combinations of prebiotics andprobiotics) (see, e.g., WO 2011/022542). Autoimmune and inflammatorydiseases are characterized by an inappropriate immunological intoleranceor an abnormal immune response, and affect up to 50 million Americans.Current treatments for such conditions, such as immunosuppressant drugs,carry a risk of dangerous systemic side effects such as infection, organdamage, and the development of new autoimmunities. There is therefore aneed for improved diagnostic and prognostic measures, preventativemeasures, and treatments for autoimmune and inflammatory diseases.

A healthy microbiota provides the host with multiple benefits, includingcolonization resistance to a broad spectrum of pathogens, essentialnutrient biosynthesis and absorption, and immune stimulation thatmaintains a healthy gut epithelium and an appropriately controlledsystemic immunity. In settings of ‘dysbiosis’ or disrupted symbiosis,microbiota functions can be lost or deranged, resulting in increasedsusceptibility to pathogens, altered metabolic profiles, or induction ofproinflammatory signals that can result in local or systemicinflammation or autoimmunity. Thus, the intestinal microbiota plays asignificant role in the pathogenesis of many diseases and disorders,including a variety of pathogenic infections distal to thegastrointestinal tract. Therefore, in response to the need for durable,efficient, and effective compositions and methods for treatment ofimmune and inflammatory diseases by way of restoring or enhancingmicrobiota functions, the present invention provides compositions andmethods for treatment and prevention of immune and inflammatoryconditions associated with dysbiosis, including dysbiosis distal to thegastrointestinal tract.

SUMMARY OF THE INVENTION

Disclosed herein are therapeutic compositions containing probiotic,non-pathogenic bacterial populations and networks thereof, for theprevention, control, and treatment of diseases, disorders andconditions, in particular immune and inflammatory diseases. In someembodiments, the therapeutic compositions contain prebiotics, e.g.,carbohydrates, in conjunction with microbial populations and/or networksthereof. These compositions are advantageous in being suitable for safeadministration to humans and other mammalian subjects and areefficacious in numerous dysbiotic diseases, disorders and conditions,such as immune and inflammatory disease.

In one aspect, the invention provides a pharmaceutical compositioncomprising an isolated population of anti-inflammatory bacterial cellsof the order Clostridiales capable of decreasing the secretion of apro-inflammatory cytokine and/or increasing the secretion of ananti-inflammatory cytokine by a population of human peripheral bloodmononuclear cells (PBMCs), and a pharmaceutically acceptable excipient.In one embodiment, the secretion of a pro-inflammatory cytokine by apopulation of PBMCs is induced by Enterococcus faecalis.

In one embodiment, the anti-inflammatory bacterial cells are of thefamily Lachnospiraceae. In another embodiments, the anti-inflammatorybacterial cells are of the genus Blautia, Clostridium, Eubacterium, orRuminococcus. In one the anti-inflammatory bacterial cells are of thegenus Blautia. In another embodiment, the anti-inflammatory bacterialcells are of a species selected from the group consisting of Blautiacoccoides, Blautia faecis, Blautia glucerasea, Blautia hansenii, Blautiahyrogenotrophica, Blautia luti, Blautia obeum, Blautia producta, Blautiaschinkii, Blautia sp. M25, Blautia stercoris, Blautia wexlerae, Blautiauncultured bacterium clone BKLE_a03_2, Blautia uncultured bacteriumclone SJTU_B_14_30, Blautia uncultured bacterium clone SJTU_C_14_16,Blautia uncultured bacterium clone S1-5, and Blautia unculturedPACO000178_s. In one embodiment, the anti-inflammatory bacterial cellsare of the species Ruminococcus gnavus. In another embodiment, theanti-inflammatory bacterial cells are of the species Eubacteriumrectale.

In one embodiment, the anti-inflammatory bacterial cells comprise abacterial cell in vegetative form. In another embodiment, theanti-inflammatory bacterial cells comprise a bacterial cell in sporeform.

In one embodiment, the isolated population of anti-inflammatorybacterial cells further comprises a bacterial cell belonging to abacterial strain set forth in Table 1, Table 1A, Table 1B, Table 1C,Table 1D, Table 1E, or Table 1F.

In one embodiment, the pharmaceutical composition comprises a prebiotic.In one embodiment, the prebiotic comprises a monomer or polymer selectedfrom the group consisting of arabinoxylan, xylose, soluble fiberdextran, soluble corn fiber, polydextrose, lactose,N-acetyl-lactosamine, glucose, and combinations thereof. In anotherembodiment, the prebiotic comprises a monomer or polymer selected fromthe group consisting of galactose, fructose, rhamnose, mannose, uronicacids, 3′-fucosyllactose, 3′-sialylactose, 6′-sialyllactose,lacto-N-neotetraose, 2′-2′-fucosyllactose, and combinations thereof. Inone embodiment, the prebiotic comprises a monosaccharide selected fromthe group consisting of arabinose, fructose, fucose, galactose, glucose,mannose, D-xylose, xylitol, ribose, and combinations thereof. In anotherembodiment, the prebiotic comprises a disaccharide selected from thegroup consisting of xylobiose, sucrose, maltose, lactose, lactulose,trehalose, cellobiose, and combinations thereof. In yet anotherembodiment, the prebiotic comprises a polysaccharide, wherein thepolysaccharide is xylooligosaccharide.

In one embodiment, the prebiotic comprises a sugar selected from thegroup consisting of arabinose, fructose, fucose, lactose, galactose,glucose, mannose, D-xylose, xylitol, ribose, xylobiose, sucrose,maltose, lactose, lactulose, trehalose, cellobiose, xylooligosaccharide,and combinations thereof. In one embodiment, the sugar is xylose.

In one embodiment, the pro-inflammatory cytokine is selected from thegroup consisting of IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1, MIP1α,MIP1β, TNFα, and combinations thereof.

In one embodiment, the anti-inflammatory cytokine is selected from thegroup consisting of IL-10, IL-13, IL-4, IL-5, TGFβ, and combinationsthereof.

In one embodiment, the pharmaceutical composition is formulated for oraladministration. In another embodiment, the pharmaceutical composition isformulated for rectal administration.

In one embodiment, the anti-inflammatory bacterial cells decrease thesecretion of a pro-inflammatory cytokine and/or increase the secretionof an anti-inflammatory cytokine by a population of human peripheralblood mononuclear cells (PBMCs) in vitro.

In another aspect, the invention provides a method for reducinginflammation in a subject, the method comprising administering apharmaceutical composition of the invention to thereby reduceinflammation in the subject.

In one embodiment, the subject has an autoimmune or inflammatorydisorder. In one embodiment, the autoimmune or inflammatory disorder isselected from the group consisting of graft-versus-host disease (GVHD),an inflammatory bowel disease (IBD), ulterative colitis, Crohn'sdisease, multiple sclerosis (MS), systemic lupus erythematosus (SLE),type I diabetes, rheumatoid arthritis, Sjögren's syndrome, and Celiacdisease.

In one embodiment, the pharmaceutical composition is administeredorally. In another embodiment, the pharmaceutical composition isadministered rectally.

In one embodiment, administration of the pharmaceutical compositionreduces inflammation in the gastrointestinal tract of the subject. Inanother embodiment, administration of the pharmaceutical compositionreduces inflammation at a site distal to the gastrointestinal tract ofthe subject. In one embodiment, the distal site is the placenta, thespleen, the skin the liver, the uterus, the blood, an eye/conjunctiva,the mouth an ear, the nose, a lung, the liver, the pancreas, the brain,the embryonic sac, or vagina of the subject. In another embodiment, thedistal site is the circulatory system, the reproductive tract, thecardiovascular system, the nervous system, or a combination thereof.

In one embodiment, the subject has a dysbiosis. In one embodiment, thedysbiosis is a gastrointestinal dysbiosis. In another embodiment, thedysbiosis is a distal dysbiosis.

In one embodiment, the anti-inflammatory bacterial cells of thepharmaceutical composition engraft in the gastrointestinal tract of thesubject.

In one embodiment, the method further comprises administering aprebiotic to the subject.

BRIEF DESCRIPTION OF THE TABLES

Table 1 provides a list of Operational Taxonomic Units (OTU) withtaxonomic assignments made to Genus, Species, and Phylogenetic Clade.Clade membership of bacterial OTUs is based on 16S sequence data. Cladesare defined based on the topology of a phylogenetic tree that isconstructed from full-length 16S sequences using maximum likelihoodmethods familiar to individuals with ordinary skill in the art ofphylogenetics. Clades are constructed to ensure that all OTUs in a givenclade are: (i) within a specified number of bootstrap supported nodesfrom one another, and (ii) within 5% genetic similarity. OTUs that arewithin the same clade can be distinguished as genetically andphylogenetically distinct from OTUs in a different clade based on 16S-V4sequence data, while OTUs falling within the same clade are closelyrelated. OTUs falling within the same clade are evolutionarily closelyrelated and may or may not be distinguishable from one another using16S-V4 sequence data. Members of the same clade, due to theirevolutionary relatedness, play similar functional roles in a microbialecology such as that found in the human gut. Compositions substitutingone species with another from the same clade are likely to haveconserved ecological function and therefore are useful in the presentinvention. All OTUs are denoted as to their putative capacity to formspores and whether they are a Pathogen or Pathobiont (see Definitionsfor description of “Pathobiont”). NIAID Priority Pathogens are denotedas ‘Category-A’, ‘Category-B’, or ‘Category-C’, and OpportunisticPathogens are denoted as ‘OP’. OTUs that are not pathogenic or for whichtheir ability to exist as a pathogen is unknown are denoted as ‘N’. The‘SEQ ID Number’ denotes the identifier of the OTU in the SequenceListing File and ‘Public DB Accession’denotes the identifier of the OTUin a public sequence repository. See, e.g., WO 2014/121304.

Table 1A provides a list of exemplary bacteria useful in the presentinvention.

Table 1B provides a list of exemplary bacteria useful in the presentinvention.

Table 1C provides a list of exemplary bacteria useful in the presentinvention.

Table 1D provides a list of exemplary bacteria useful in the presentinvention.

Table 1E provides a list of exemplary bacteria useful in the presentinvention. These bacteria are preferably down-modulated in a subject.

Table 1F provides a list of exemplary bacteria that may be used in theinvention. These bacteria are preferably up-modulated in a subject.

Table 2A lists species identified as “germinable” and “sporulatable” bycolony picking approach.

Table 2B lists species identified as “germinable” using 16S colonypicking approach.

Table 2C lists species identified as “sporulatable” using 16s-V4 NGSapproach. See, e.g., WO 2014/121304.

Table 3 lists anaerobic bacterial species tested for carbon source usage(Biolog plates).

Table 4 lists exemplary prebiotics/carbon sources.

Table 5 provides bacterial species detected at low frequency in vaginalsamples from vancomycin-treated mice (day 6) that were not present inuntreated mice (day 0).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph depicting serum endotoxin levels (EU/ml) over timefollowing treatment with xylose. Treatment of mice with xylose alonereduces basal levels of serum endotoxin (day 14 vs day 0). Antibiotictreatment (Ciprofloxacin (cipro) or enrofloxacin (enro)) leads to anincrease in serum endotoxin levels (measured 2 days after a 5 daycourse, at day 0) with a return to baseline by day 14. Xylosecounteracts the endotoxin increase caused by cipro but not enroantibiotic treatment.

FIG. 2 (a-o) is a panel of graphs showing the time course of Th1 relatedcytokines that were released by human peripheral mononuclear cells(PBMCs) incubated with Ruminococcus gnavus (Epv 1), Eubacterium rectale(Epv 2), Blautia luti (Epv 3), Blautia wexlerae (Epv 5) and Enterococcusfaecalis (Epv 8), or combinations of each bacterium with E. faecalis.Amounts of interferon gamma (IFN-γ), IL-12p70, IL-6, IL-2 and TNFα thatwere released in culture supernatants by PBMCs were measured after 24,48 and 72 hours. a) IFN-γ concentration (pg/ml) after 24 hours. b) IFN-γconcentration (pg/ml) after 48 hours. c) IFN-γ concentration (pg/ml)after 72 hours. d) IL-12p70 concentration (pg/ml) after 24 hours. e)IL-12p70 concentration (pg/ml) after 48 hours. f) IL-12p70 concentration(pg/ml) after 72 hours. g) IL-6 concentration (pg/ml) after 24 hours. h)IL-6 concentration (pg/ml) after 48 hours. i) IL-6 concentration (pg/ml)after 72 hours. j) IL-2 concentration (pg/ml) after 24 hours. k) IL-2concentration (pg/ml) after 48 hours. l) IL-2 concentration (pg/ml)after 72 hours. m) TNFα concentration (pg/ml) after 24 hours. n) TNFαconcentration (pg/ml) after 48 hours. o) TNFα concentration (pg/ml)after 72 hours.

FIG. 3 (a-i) is a panel of graphs showing the time course of Th2 relatedcytokines that were released by human PBMCs incubated with R. gnavus(Epv 1), E. rectale (Epv 2), B. luti (Epv 3), B. wexlerae (Epv 5) and E.faecalis (Epv 8), or combinations of each bacterium with E. faecalis.Amounts of IL-13, IL-4 and IL-5 that were released in culturesupernatants by PBMCs were measured after 24, 48 and 72 hours. a) IL-13concentration (pg/ml) after 24 hours. b) IL-13 concentration (pg/ml)after 48 hours. c) IL-13 concentration (pg/ml) after 72 hours. d) IL-4concentration (pg/ml) after 24 hours. e) IL-4 concentration (pg/ml)after 48 hours. f) IL-4 concentration (pg/ml) after 72 hours. g) IL-5concentration (pg/ml) after 24 hours. h) IL-5 concentration (pg/ml)after 48 hours. i) IL-5 concentration (pg/ml) after 72 hours.

FIG. 4 (a-i) is a panel of graphs showing the time course of Th9, Th17and Treg cytokines that were released by human PBMCs incubated with R.gnavus (Epv 1), E. rectale (Epv 2), B. luti (Epv 3), B. wexlerae (Epv 5)and E. faecalis (Epv 8), or combinations of each bacterium with E.faecalis. Amounts of IL-9, IL-17 and IL-10 that were released in culturesupernatants by PBMCs were measured after 24, 48 and 72 hours. a) IL-9concentration (pg/ml) after 24 hours. b) IL-9 concentration (pg/ml)after 48 hours. c) IL-9 concentration (pg/ml) after 72 hours. d) IL-17concentration (pg/ml) after 24 hours. e) IL-17 concentration (pg/ml)after 48 hours. f) IL-17 concentration (pg/ml) after 72 hours. g) IL-10concentration (pg/ml) after 24 hours. h) IL-10 concentration (pg/ml)after 48 hours. i) IL-10 concentration (pg/ml) after 72 hours.

FIG. 5 (a-x) is a panel of graphs showing the time course of monocyte,macrophage and neutrophil-derived inflammatory cytokines that werereleased by human PBMCs incubated with R. gnavus (Epv 1), E. rectale(Epv 2), B. luti (Epv 3), B. wexlerae (Epv 5) and E. faecalis (Epv 8),or combinations of each bacterium with E. faecalis. Amounts of monocytechemotactic protein 1 (MCP-1), macrophage inflammatory protein 1β(MIP1β), macrophage inflammatory protein 1α (MIP1α), regulated onactivation, normal T expressed and secreted protein (RANTES),interleukin-1α (IL-1α), interleukin-1β (IL1β), interferon α2 (IFN-α2)and interleukin-8 (IL-8) that were released in culture supernatants byPBMCs were measured after 24, 48 and 72 hours. a) MCP-1 concentration(pg/ml) after 24 hours. b) MCP-1 concentration (pg/ml) after 48 hours.c) MCP-1 concentration (pg/ml) after 72 hours. d) MIP10 concentration(pg/ml) after 24 hours. e) MIP10 concentration (pg/ml) after 48 hours.f) MIP10 concentration (pg/ml) after 72 hours. g) MIP1α concentration(pg/ml) after 24 hours. h) MIP1a concentration (pg/ml) after 48 hours.i) MIP1a concentration (pg/ml) after 72 hours. j) RANTES concentration(pg/ml) after 24 hours. k) RANTES concentration (pg/ml) after 48 hours.l) RANTES concentration (pg/ml) after 72 hours. m) IL-1α concentration(pg/ml) after 24 hours. n) IL-1α concentration (pg/ml) after 48 hours.o) IL-1α concentration (pg/ml) after 72 hours. p) IL1β concentration(pg/ml) after 24 hours. q) IL1β concentration (pg/ml) after 48 hours. r)IL1β concentration (pg/ml) after 72 hours. s) IFN-α2 concentration(pg/ml) after 24 hours. t) IFN-α2 concentration (pg/ml) after 48 hours.u) IFN-α2 concentration (pg/ml) after 72 hours. v) IL-8 concentration(pg/ml) after 24 hours. w) IL-8 concentration (pg/ml) after 48 hours. x)IL-8 concentration (pg/ml) after 72 hours.

FIG. 6 (a-d) is a panel of graphs showing the secreted levels ofcytokines IFNγ (Ifng), IL-12p70, IL-1α (IL-1α), IL-6, IL-8, MCP1, MIP1α(MIP1α), MIP1β (MIP1b), TNFα (TNFα), IL-10, IL-13, IL-9, IL-4, IL-5,IL-17α (IL-17A) and IL-2 produced by PBMCs in the presence of a) R.gnavus, b) B. wexlerae, c) E. rectale and d) B. luti, alone or incombination with E. faecalis (Epv 8), relative to levels secretedfollowing treatment with E. faecalis alone for 24 hours (E.faecalis=100%).

FIG. 7 (a-p) is a panel of graphs that show the effect of R. gnavus(Epv1) on cytokine concentration (pg/ml) either alone or in combinationwith Epv 8 (E. faecalis) on cytokine production by human PBMCs (pg/ml).a) IL-6, b) IFN-γ, c) IL-13, d) IL-10, e) IL-12p70, f) MCP-1, g) IL-8,h) IL17A, i) IL-α, j) IL-9, k) IL-2, l) IL-4, m) IL-5, n) MIP-1a, o)MIP-1β, p) TNF-α.

FIG. 8 (a-p) is a panel of graphs that show the effect of E. rectale(Epv2) on cytokine concentration (pg/ml) either alone or in combinationwith Epv 8 (E. faecalis) on cytokine production by human PBMCs (pg/ml).a) IL-6, b) IFN-γ, c) IL-13, d) IL-10, e) IL-12p70, f) MCP-1, g) IL-8,h) IL17A, i) IL-α, j) IL-9, k) IL-2, l) IL-4, m) IL-5, n) MIP-1a, o)MIP-1β, p) TNF-α.

FIG. 9 (a-p) is a panel of graphs that show the effect of B. luti (Epv3)on cytokine concentration (pg/ml) either alone or in combination withEpv 8 (E. faecalis) on cytokine production by human PBMCs (pg/ml). a)IL-6, b) IFN-γ, c) IL-13, d) IL-10, e) IL-12p70, f) MCP-1, g) IL-8, h)IL17α, i) IL-α, j) IL-9, k) IL-2, l) IL-4, m) IL-5, n) MIP-1α, o)MIP-1β, p) TNF-α.

FIG. 10 (a-p) is a panel of graphs that show the effect of B. wexlarae)on cytokine concentration (pg/ml) either alone or in combination withEpv 8 (E. faecalis) on cytokine production by human PBMCs (pg/ml). a)IL-6, b) IFN-γ, c) IL-13, d) IL-10, e) IL-12p70, f) MCP-1, g) IL-8, h)IL17α, i) IL-α, j) IL-9, k) IL-2, l) IL-4, m) IL-5, n) MIP-1α, o)MIP-1β, p) TNF-α.

FIG. 11 (a-d) is a panel of graphs showing that (a-b) EPV3 is capable ofinducing a desirable anti-inflammatory cytokine profile for treating orpreventing GVHD and (c-d) EPV5 induces a suboptimal profile for GVHD.

FIG. 12 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv6 (Clostridiumleptum).

FIG. 13 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv15 (Blautiafaecis).

FIG. 14 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv20(Blautia/Ruminococcus obeum ATCC 29174).

FIG. 15 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv21 (Blautiaproducta ATCC 27340).

FIG. 16 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv22 (Blautiacoccoides ATCC 29236).

FIG. 17 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv23 (Blautiahydrogenotrophica ATCC BAA-2371).

FIG. 18 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv24 (Blautiahansenii ATCC27752).

FIG. 19 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv35 (Eubacteriumrectale).

FIG. 20 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv47 (previouslyuncultured Blautia, similar to GQ898099_s S1-5).

FIG. 21 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv51 (previouslyuncultured Blautia, similar to SJTU_C_14_16).

FIG. 22 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv52 (Blautiawexlerae (SJTU_B_09_77)).

FIG. 23 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv54 (Blautia lutiELU0087-T13-S-NI_000247).

FIG. 24 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv64 (Blautiawexlerae WAL 14507).

FIG. 25 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv78 (Blautia obeum).

FIG. 26 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv102 (Ruminococcusgnavus).

FIG. 27 (a-b) depicts the production of (a) pro-inflammatory (IL-12p70,IFNγ, IP-10, IL-1RA) and (b) anti-inflammatory (IL-10, IL-4, IL-13)cytokines by human PBMCs following treatment with Epv114 (Blautia luti(BlnIX)).

FIG. 28 (a-d) presents results from flow cytometry analysis of T cellpopulations in human PBMCs incubated in the presence of variouscommensal bacteria, determined using flow cytometry. A) Proportion ofTreg cells (CD25⁺CD127^(lo)); B) Proportion of Th17 cells (CXCR3⁻CCR6⁺); C) Proportion of Th1 cells (CXCR3⁺CCR6⁻); D) Proportion of Th2cells (CXCR3⁻ CCR6⁻). Bacterial strains are as follows: Epv 1: R.gnavus; Epv 3: B. luti; Epv 2: E. rectale; Epv 5: B. wexlerae; Epv. 8:E. faecalis; Epv 20: B. obeum; Epv 21: B. producta; Epv 24: B. hansenii.The results are shown as percent (%) of CD3ε⁺CD4⁺ cells.

FIG. 29 (a-u) presents the preferred carbon sources utilized by variouscommensal bacteria. (a) R. gnavus; (b) E. rectale; (c) C. leptum; (d) B.luti; (e) B. wexlerae; (f) B. faecis; (g) B. obeum; (h) B. producta; (i)B. coccoides; (j) B. hydrogenotrophica; (k) B. hansenii; (1) B. lutiBlnl X; (m) B. luti ELU; (n) R. gnavus; (o) B. faecis; (p) R. torques;(q) B. wexlerae WAL14507; (r) B. wexlerae SJTU; (s) SJTU1416; (t)GQ8980099; (u) E. rectale.

FIG. 30 graphically depicts levels of serum IFNγ before, during, andafter treatment with a prebiotic formulation containing xylose.

FIG. 31 is a graph that shows the change in Chao1 diversity (indicatorof community richness) over time in subjects administered xylose threetimes per day (TID) at 1, 2, 8, 12.5 or 15 grams.

FIG. 32 depicts the impact of oral vancomycin on the microbiome of thegut and the vagina, by principal component analysis (PCA).

DETAILED DESCRIPTION

Disclosed herein are therapeutic compositions (e.g., pharmaceuticalcompositions) containing bacterial entities (e.g., anti-inflammatorybacterial cells) and optionally containing a prebiotic for theprevention, control, and treatment of immune and inflammatory diseases,disorders and conditions. These compositions are advantageous in beingsuitable for safe administration to humans and other mammalian subjectsand are efficacious in treating or preventing numerous immune andinflammatory diseases and gastrointestinal diseases, disorders andconditions associated with a dysbiosis.

The microbes that inhabit the human gastrointestinal tract, skin, lungs,vagina, and other niches are starting to be understood and appreciatedfor their roles in human health and disease (see, e.g., Human MicrobiomeProject Consortium (2012) NATURE 486(7402): 207-14). Aspects of theinvention are based, in part, on the realization that, althoughautoimmune and inflammatory diseases are often attributed to geneticmutations, these conditions are also influenced by microbes. It is alsoappreciated that, because microbes not only interact with the host butwith one another, the immunomodulatory behavior of microbes can dependon relationships between microbes. For example, a microbial network in agiven niche may comprise diverse microbes that all accomplish one ormore of the same functions, or may instead comprise diverse microbesthat all individually contribute to accomplish one or more functions.For example, microbes in a given niche may influence and/or regulate theimmunomodulatory behavior of other microbes in the same niche, or in adistal niche. In another example, microbes in a given niche may competewith one another for nutrients or space.

Microbes may influence the risk, progression, or treatment efficacy ofan autoimmune or inflammatory disease. In certain aspects, microbes playa role in the prevention of an autoimmune or inflammatory disease or inthe suppression of an innate or adaptive immune response. Microbes mayalso stimulate an inflammatory immune response to contribute to,increase the risk of, or worsen the symptoms of an autoimmune orinflammatory disease. Some microbes may be associated with lower diseaseseverity or mortality.

Also disclosed herein are compositions and methods for the preventionand/or treatment of autoimmune and inflammatory diseases in humansubjects.

Definitions

As used in the specification and the appended claims, the singular forms“a,” “an,” and “the” include plural referents unless the context clearlydictates otherwise. Thus, for example, “a compound” includes mixtures ofcompounds.

The term “about” or “approximately” means within an acceptable errorrange for the particular value as determined by one of ordinary skill inthe art, which will depend in part on how the value is measured ordetermined, i.e., the limitations of the measurement system. Forexample, “about” can mean within 3 or more than 3 standard deviations,per the practice in the art. Alternatively, “about” can mean a range ofup to 20%, or up to 10%, or up to 5%, or up to 1% of a given value.Alternatively, particularly with respect to biological systems orprocesses, the term can mean within an order of magnitude, or within5-fold, or within 2-fold, of a value.

As used herein, the term “purified bacterial preparation” refers to apreparation that includes “isolated” bacteria or bacteria that have beenseparated from at least one associated substance found in a sourcematerial or any material associated with the bacteria in any processused to produce the preparation.

A “bacterial entity” includes one or more bacteria. Generally, a firstbacterial entity is distinguishable from a second bacterial entity.

As used herein, the term “formation” refers to synthesis or production.

As used herein, the term “inducing” means increasing the amount oractivity of a given material as dictated by context.

As used herein, the term “depletion” refers to reduction in amount of.

As used herein, a “prebiotic” refers to an ingredient that allowsspecific changes both in the composition and/or activity in thegastrointestinal microbiota that may (or may not) confer benefits uponthe host. In some embodiments, a prebiotic can be a comestible food orbeverage or ingredient thereof. Prebiotics may include complexcarbohydrates, amino acids, peptides, minerals, or other essentialnutritional components for the survival of the bacterial composition.Prebiotics include, but are not limited to, amino acids, biotin,fructooligosaccharide, galactooligosaccharides, hemicelluloses (e.g.,arabinoxylan, xylan, xyloglucan, and glucomannan), inulin, chitin,lactulose, mannan oligosaccharides, oligofructose-enriched inulin, gums(e.g., guar gum, gum arabic and carregenaan), oligofructose,oligodextrose, tagatose, resistant maltodextrins (e.g., resistantstarch), trans-galactooligosaccharide, pectins (e.g., xylogalactouronan,citrus pectin, apple pectin, and rhamnogalacturonan-I), dietary fibers(e.g., soy fiber, sugarbeet fiber, pea fiber, corn bran, and oat fiber)and xylooligosaccharides.

As used herein, “predetermined ratios” refer to ratios determined orselected in advance.

As used herein, “germinable bacterial spores” are spores capable offorming vegetative cells in response to a particular cue (e.g., anenvironmental condition or a small molecule).

As used herein, “detectably present” refers to presence in an amountthat can be detected using assays provided herein or otherwise known inthe art that exist as of the filing date.

As used herein, “augmented” refers to an increase in amount and/orlocalization within to a point where it becomes detectably present.

As used herein, “fecal material” refers to a solid waste product ofdigested food and includes feces or bowel washes.

As used herein, the phrase “host cell response” refers to a responseproduced by a cell of a host organism.

As used herein, a “mammalian subject protein” refers to a proteinproduced by a mammalian subject and encoded by the mammalian subjectgenome. The term mammalian subject protein includes proteins that havebeen post-translationally processed and/or modified.

As used herein, the term “food-derived” refers to a protein orcarbohydrate found in a consumed food.

As used herein, the term “biological material” refers to a materialproduced by a biological organism.

As used herein, the term “detection moiety” refers to an assay componentthat functions to detect an analyte.

As used herein, the term “incomplete network” refers to a partialnetwork that lacks at least one of the entire set of components neededto carry out one or more network functions.

As used herein, the term “supplemental” refers to something that isadditional and non-identical.

As used herein, a composition is “substantially free” of microbes whenmicrobes are absent or undetectable as determined by the use of standardgenomic and microbiological techniques. A composition is “substantiallyfree” of a prebiotic or immunostimulatory carbohydrate whennon-microbial carbohydrates are absent or undetectable as determined bythe use of standard biochemical techniques (e.g., dye-based assays).

Microbial agents (individual or populations of microbes, microbialnetworks or parts of networks, or microbial metabolites) are consideredto be “exogenous” to a subject (e.g., a human or non-human animal), acell, tissue, organ or other environment of a human or non-human animal,if said subject, or said cell, tissue, organ or other environment of thesubject, does not contain detectable levels of the microbial agent.

A microbial agent or population thereof is “heterologous” or“heterologously contained” on or in a host environment when, e.g., themicrobial agent or population is administered or disposed on, or in thehost, or host environment in a number, concentration, form or othermodality that is not found in the host prior to administration of themicrobial agent or population, or when the microbial agent or populationcontains an activity or structural component different from a host thatdoes not naturally have the microbial agent within the targetenvironment to which the microbe is administered or thereafter disposed.

As used herein, the term “antioxidant” is understood to include any oneor more of various substances such as beta-carotene (a vitamin Aprecursor), vitamin C, vitamin E, and selenium) that inhibit oxidationor reactions promoted by Reactive Oxygen Species (“ROS”) and otherradical and non-radical species. Additionally, antioxidants aremolecules capable of slowing or preventing the oxidation of othermolecules. Non-limiting examples of antioxidants include astaxanthin,carotenoids, coenzyme Q10 (“CoQ10”), flavonoids, glutathione, Goji(wolfberry), hesperidin, lactowolfberry, lignan, lutein, lycopene,polyphenols, selenium, vitamin A, vitamin C, vitamin E, zeaxanthin, orcombinations thereof.

“Backbone network ecology” or simply “backbone network” or “backbone”are compositions of microbes that form a foundational composition thatcan be built upon or subtracted from to optimize a network ecology orfunctional network ecology to have specific biological characteristicsor to comprise desired functional properties, respectively. Microbiometherapeutics can be comprised of these “backbone networks ecologies” intheir entirety, or the “backbone networks” can be modified by theaddition or subtraction of “R-groups” to give the network ecologydesired characteristics and properties. “R-groups” can be defined inmultiple terms including, but not limited to: individual OTUs,individual or multiple OTUs derived from a specific phylogenetic cladeor a desired phenotype such as the ability to form spores, or functionalbacterial compositions that comprise. “Backbone networks” can comprise acomputationally derived network ecology in its entirety, or can comprisesubsets of the computationally-derived network ecology that representkey nodes in the network that contribute to efficacy such as but notlimited to a composition of Keystone OTUs. The number of organisms in ahuman gastrointestinal tract, is indicative of the functional redundancyof a healthy gut microbiome ecology (see, e.g., The Human MicrobiomeConsortia (2012). This redundancy makes it highly likely thatnon-obvious subsets of OTUs or functional pathways (i.e., “backbonenetworks”) are critical to maintaining states of health and/orcatalyzing a shift from a dysbiotic state to one of health. One way ofexploiting this redundancy is through the substitution of OTUs thatshare a given clade (see below) or by adding members of a clade notfound in the backbone network.

“Bacterial composition” refers to a consortium of microbes comprisingtwo or more OTUs. Backbone network ecologies, functional networkecologies, network classes, and core ecologies are all types ofbacterial compositions. As used herein, bacterial composition includes atherapeutic microbial composition, a prophylactic microbial composition,a spore population, a purified spore population, or an ethanol treatedspore population.

“Bacterial translocation” refers to the passage of one or more bacteriaacross the epithelial layer of any organ of a human or non-human animal.

“Network ecology” refers to a consortium of clades or OTUs that co-occurin some number of subjects. As used herein, a “network” is definedmathematically by a graph delineating how specific nodes (i.e., cladesor OTUs) and edges (connections between specific clades or OTUs) relateto one another to define the structural ecology of a consortium ofclades or OTUs. Any given network ecology will possess inherentphylogenetic diversity and functional properties.

A network ecology can also be defined in terms of its functionalcapabilities where for example the nodes would be comprised of elementssuch as, but not limited to, enzymes, clusters of orthologous groups. orKEGG Orthology Pathways; these networks are referred to as a “functionalnetwork ecology”. Functional network ecologies can be reduced topractice by defining the group of OTUs that together comprise thefunctions defined by the functional network ecology.

The terms “network class”, “core network” and “network class ecology”refer to a group of network ecologies that in general arecomputationally determined to comprise ecologies with similarphylogenetic and/or functional characteristics. A network classtherefore contains important biological features, defined eitherphylogenetically or functionally, of a group (i.e., a cluster) ofrelated network ecologies. One representation of a core network ecologyis a designed consortium of microbes, typically non-pathogenic bacteria,that represents core features of a set of phylogenetically orfunctionally related network ecologies seen in many different subjects.In many occurrences, a core network, while designed as described herein,exists as a network ecology observed in one or more subjects. Corenetwork ecologies are useful for reversing or reducing a dysbiosis insubjects where the underlying, related network ecology has beendisrupted.

“Bacterial translocation” refers to the passage of one or more bacteriaacross the epithelial layer of any organ of a human or non-human animal.

“Clade” refers to the OTUs or members of a phylogenetic tree that aredownstream of a statistically valid node in a phylogenetic tree. Theclade comprises a set of terminal leaves in the phylogenetic tree (i.e.,tips of the tree) that are a distinct monophyletic evolutionary unit andthat share some extent of sequence similarity. Clades are hierarchical,in one embodiment, the node in a phylogenetic tree that is selected todefine a clade is dependent on the level of resolution suitable for theunderlying data used to compute the tree topology.

The “colonization” of a host organism includes the non-transitoryresidence of a bacterium or other microscopic organism. As used herein,“reducing colonization” of a host subject's gastrointestinal tract orvagina (or any other microbiota niche) by a pathogenic or non-pathogenicbacterium includes a reduction in the residence time of the bacterium inthe gastrointestinal tract or vagina, as well as a reduction in thenumber (or concentration) of the bacterium in the gastrointestinal tractor vagina, or adhered to the luminal surface of the gastrointestinaltract. The reduction in colonization can be permanent or occur during atransient period of time. Measuring reductions of adherent pathogens canbe demonstrated directly, e.g. by determining pathogenic burden in abiopsy sample, or reductions may be measured indirectly, e.g., bymeasuring the pathogenic burden in the stool of a mammalian host.

A “combination” of two or more bacteria includes the physicalco-existence of the two bacteria, either in the same material or productor in physically connected products, as well as the temporalco-administration or co-localization of the two bacteria.

“Cytotoxic” activity of bacterium includes the ability of a bacteriumkill a cell (e.g., a host cell or a bacterial cell). A “cytostatic”activity of a bacterium includes the ability to inhibit (e.g., partiallyor fully) the growth, metabolism, and/or proliferation of a cell (e.g.,a bacterial cell or a host cell).

“Dysbiosis” refers to a state of the microbiota or microbiome of the gutor other body area, including mucosal or skin surfaces (or any othermicrobiota niche) in which the normal diversity and/or function of theecological network is disrupted. Any disruption from the preferred(e.g., ideal) state of the microbiota can be considered a dysbiosis,even if such dysbiosis does not result in a detectable decrease inhealth. This state of dysbiosis may be unhealthy (e.g., result in adiseased state), it may be unhealthy under only certain conditions, orit may prevent a subject from becoming healthier. Dysbiosis may be dueto a decrease in diversity of the microbiota population composition, theovergrowth of one or more population of pathogens (e.g., a population ofpathogenic bacteria) or pathobionts, the presence of and/or overgrowthof symbiotic organisms able to cause disease only when certain geneticand/or environmental conditions are present in a patient, or a shift toan ecological network that no longer provides a beneficial function tothe host and therefore no longer promotes health. A state of dysbiosismay lead to a disease or disorder (e.g. a gastrointestinal disease,disorder or condition), or the state of dysbiosis may lead to a diseaseor disorder (e.g., a gastrointestinal disease, disorder or condition)only under certain conditions, or the state of dysbiosis may prevent asubject from responding to treatment or recovering from a disease ordisorder (e.g., a gastrointestinal disease, disorder or condition).

The term “distal” generally is used in relation to the gastrointestinaltract, specifically the intestinal lumen, of a human or other mammal.Thus, a “distal dysbiosis” includes a dysbiosis outside of the lumen ofthe gastrointestinal tract, and a “distal microbiota” includes amicrobiota outside of the lumen of the gastrointestinal tract. Inspecified instances, the term “distal” may be used in relation to thesite of administration, engraftment, or colonization of a composition,e.g., a probiotic composition, of the invention. For example, if aprobiotic composition is administered vaginally, a “distal” effect ofthe composition would occur outside the vagina.

“Gastrointestinal dysbiosis” refers to a state of the microbiota ormicrobiome of the gut in which the normal diversity and/or function ofthe ecological network or niche is disrupted. The term “gut” as usedherein is meant to refer to the entire gastrointestinal or digestivetract (also referred to as the alimentary canal) and it refers to thesystem of organs within multi-cellular animals which takes in food,digests it to extract energy and nutrients, and expels the remainingwaste. As used herein the term “gastrointestinal tract” refers to theentire digestive canal, from the oral cavity to the rectum. The term“gastrointestinal tract” includes, but is not limited to, mouth andproceeds to the esophagus, stomach, small intestine, large intestine,rectum and, finally, the anus.

“Germinant” is a material or composition, or a physical-chemicalprocess, capable of inducing the germination of vegetative bacterialcells from dormant spores, or the proliferation of vegetative bacterialcells, either directly or indirectly in a host organism and/or in vitro.

“Inhibition” of a pathogen or non-pathogen encompasses the inhibition ofany desired function or activity of the pathogen or non-pathogen by theprobiotic, e.g. bacterial, compositions of the present invention.Demonstrations of inhibition, such as a decrease in the growth of apathogenic bacterial cell population or a reduction in the level ofcolonization of a pathogenic bacterial species are provided herein andotherwise recognized by one of ordinary skill in the art. Inhibition ofa pathogenic or non-pathogenic bacterial population's “growth” mayinclude inhibiting an increase in the size of a pathogenic ornon-pathogenic bacterial cell population and/or inhibiting theproliferation (or multiplication) of a pathogenic or non-pathogenicbacterial cell population. Inhibition of colonization of a pathogenic ornon-pathogenic bacterial species may be demonstrated by measuring andcomparing the amount or burden of the bacterial species before and aftera treatment. An “inhibition” or the act of “inhibiting” includes thetotal cessation and partial reduction of one or more activities of apathogen, such as growth, proliferation, colonization, and function. Asused herein, inhibition includes cytostatic and/or cytotoxic activities.Inhibition of function includes, for example, the inhibition ofexpression of a pathogenic gene product (e.g., the genes encoding atoxin and/or toxin biosynthetic pathway, or the genes encoding astructure required for intracellular invasion (e.g., an invasive pilus))induced by the bacterial composition.

“Isolated” encompasses a bacterium or other entity or substance (e.g., abacterial population or a prebiotic) that has been (1) separated from atleast some of the components with which it was associated when initiallyproduced (whether in nature or in an experimental setting), and/or (2)produced, prepared, purified, and/or manufactured by the hand of man.Isolated bacteria includes, for example, those bacteria that arecultured, even if such cultures are not monocultures. Isolated bacteriamay be separated from at least about 10%, about 20%, about 30%. about40%. about 50%, about 60%, about 70%, about 80%, about 90%, or more ofthe other components with which they were initially associated. In someembodiments, isolated bacteria are more than about 80%, about 85%, about90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%,about 97%, about 98%, about 99%, or more than about 99% pure. As usedherein, a substance is “pure” if it is substantially free of othercomponents. The terms “purify,” “purifying” and “purified” refer to abacterium or other material that has been separated from at least someof the components with which it was associated either when initiallyproduced or generated (e.g., whether in nature or in an experimentalsetting), or during any time after its initial production. A bacteriumor a bacterial population may be considered purified if it is isolatedat or after production, such as from a material or environmentcontaining the bacterium or bacterial population, or by passage throughculture. A purified bacterium or bacterial population may contain othermaterials up to about 1%, 2%. 3%, 4%. 5%, 6%, 7%, 8%, 9%, 10%, about20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%,about 90%, or above about 90%. and still be considered “isolated.” Insome embodiments, purified bacteria and bacterial populations are morethan about 80%, about 85%, about 90%, about 91%, about 92%, about 93%,about 94%, about 95%, about 96%, about 97%, about 98%, about 99%. ormore than about 99% pure. In the instance of bacterial compositionsprovided herein, the one or more bacterial types present in thecomposition can be independently purified from one or more otherbacteria produced and/or present in the material or environmentcontaining the bacterial type. In some embodiments, bacterialcompositions and the bacterial components thereof are purified fromresidual habitat products. In some embodiments, pharmaceuticalcompositions (e.g., bacterial compositions) contain a defined mixture ofisolated bacteria. For example, in some embodiments, the pharmaceuticalcomposition (e.g., probiotic composition) contains no more than 100bacterial species. For example, in some embodiments, the pharmaceuticalcomposition contains no more than 75 bacterial species. In otherembodiments, the pharmaceutical composition contains no more than 50bacterial species, e.g., no more than 40 bacterial species, no more than30 bacterial species, no more than 25 bacterial species, no more than 20bacterial species, no more than 15 bacterial species, no more than 10bacterial species, etc. In other embodiments, the pharmaceuticalcomposition contains no more than 10 bacterial species, e.g., 10bacterial species, 9 bacterial species, 8 bacterial species, 7 bacterialspecies, 6 bacterial species, 5 bacterial species, 4 bacterial species,3 bacterial species, 2 bacterial species, 1 bacterial species. In someembodiments, the pharmaceutical composition contains defined quantitiesof each bacterial species. In an exemplary embodiment, thepharmaceutical composition contains isolated bacterial populations thatare not isolated from fecal matter.

“Keystone OTU” or “keystone function” refers to one or more OTUs orfunctional pathways (e.g., KEGG or COG pathways) that are common to manynetwork ecologies or functional network ecologies and are members ofnetworks that occur in many subjects (i.e., “are pervasive). Due to theubiquitous nature of keystone OTUs and their associated functionalpathways, they are central to the function of network ecologies inhealthy subjects and are often missing, or at reduced levels, insubjects with disease. Keystone OTUs and their associated functions mayexist in low, moderate, or high abundance in subjects. A “non-keystoneOTU” or “non-keystone function” refers to an OTU or function that isobserved in a network ecology or a functional network ecology, that isnot observed in a keystone OTU or function.

“Microbiota” refers to the community of microorganisms that occur(sustainably or transiently) in and/or on a subject, (e.g, a mammal suchas a human), including, but not limited to, eukaryotes (e.g., protozoa),archaea, bacteria, and viruses (including bacterial viruses, i.e., aphage).

“Microbiome” refers to the genetic content of the communities ofmicrobes that live in and on the human body, both sustainably andtransiently, including eukaryotes, archaea, bacteria, and viruses(including, e.g., bacterial viruses (i.e., phage)), wherein “geneticcontent” includes genomic DNA, RNA such as ribosomal RNA, the epigenome,plasmids, and all other types of genetic information.

“Microbial carriage” or simply “carriage” refers to the population ofmicrobes inhabiting a niche within or on a subject (e.g., a humansubject). Carriage is often defined in terms of relative abundance. Forexample, OTU1 comprises 60% of the total microbial carriage, meaningthat OTU1 has a relative abundance of 60% compared to the other OTUs inthe sample from which the measurement is made. Carriage is most oftenbased on genomic sequencing data where the relative abundance orcarriage of a single OTU or group of OTUs is defined by the number ofsequencing reads that are assigned to that OTU/s relative to the totalnumber of sequencing reads for the sample.

“Microbial augmentation” refers to the establishment or significantincrease of a population of microbes that are (i) absent or undetectable(as determined by the use of standard genomic, biochemical and/ormicrobiological techniques) from the administered therapeutic microbialcomposition, (ii) absent, undetectable, or present at low frequencies inthe host niche (as an example: gastrointestinal tract, skin,anterior-nares, or vagina) before the delivery of the microbialcomposition, and (iii) are found, i.e., detectable, after theadministration of the microbial composition or significantly increase,for instance 2-fold, 5-fold, 1×10², 1×10³, 1×10⁴, 1×10⁵, 1×10⁶, 1×10⁷,or greater than 1×10⁸, in cases where they are present at lowfrequencies. The microbes that comprise an augmented ecology can bederived from exogenous sources, such as food and the environment, orgrow out from micro-niches within the host where they reside at lowfrequency.

The administration of the therapeutic composition (e.g., pharmaceuticalcomposition) can induce an environmental shift in the target niche thatpromotes favorable conditions for the growth of commensal microbes. Inthe absence of treatment with a therapeutic microbial composition (e.g.,a pharmaceutical composition comprising a bacterial cell population),with or without one or more prebiotics, the host can be constantlyexposed to these microbes; however, sustained growth and the positivehealth effects associated with the stable population of increased levelsof the microbes comprising the augmented ecology are not observed.

“Microbial engraftment” or simply “engraftment” refers to theestablishment of OTUs comprising a therapeutic microbial composition ina target niche. In one embodiment, the OTUs are absent in the treatedhost prior to treatment. The microbes that comprise the engraftedecology are found in the therapeutic microbial composition and establishas constituents of the host microbial ecology upon treatment. EngraftedOTUs can establish for a transient period of time, or demonstratelong-term stability in the microbial ecology that populates the hostpost-treatment with a therapeutic microbial composition. The engraftedecology can induce an environmental shift in the target niche thatpromotes favorable conditions for the growth of commensal microbescapable of catalyzing a shift from a dysbiotic ecology to onerepresentative of a healthy state.

“Ecological niche” or simply “niche” refers to the ecological space thatan organism or group of organisms (e.g., a bacterial population)occupies. Niche describes how an organism or population or organismsresponds to the distribution of resources, physical parameters (e.g.,host tissue space) and competitors (e.g., by growing when resources areabundant, and/or when predators, parasites and pathogens are scarce) andhow it in turn alters those same factors (e.g., by limiting access toresources by other organisms, acting as a food source for predatorsand/or as a consumer of prey).

“Pathobionts” or “opportunistic pathogens” refers to symbiotic organismsable to cause disease only when certain genetic and/or environmentalconditions are present in a subject.

“Phylogenetic tree” refers to a graphical representation of theevolutionary relationships of one genetic sequence to another that isgenerated using a defined set of phylogenetic reconstruction algorithms(e.g., parsimony, maximum likelihood, or Bayesian). Nodes in the treerepresent distinct ancestral sequences and the confidence of any node isprovided by a bootstrap or Bayesian posterior probability, whichmeasures branch uncertainty.

As used herein, the term “minerals” is understood to include boron,calcium, chromium, copper, iodine, iron, magnesium, manganese,molybdenum, nickel, phosphorus, potassium, selenium, silicon, tin,vanadium, zinc, or combinations thereof.

To be free of “non-comestible products” means that a bacterialcomposition or other material provided herein does not have asubstantial amount of a non-comestible product, e.g., a product ormaterial that is inedible, harmful or otherwise undesired in a productsuitable for administration, e.g., oral administration, to a humansubject. Non-comestible products are often found in preparations ofbacteria from the prior art.

“Operational taxonomic units,” “OTU” (or plural “OTUs”) refer to aterminal leaf in a phylogenetic tree and is defined by a nucleic acidsequence, e.g., the entire genome, or a specific genetic sequence, andall sequences that share sequence identity to this nucleic acid sequenceat the level of species. In some embodiments the specific geneticsequence may be the 16S sequence or a portion of the 16S sequence. Inother embodiments, the entire genomes of two entities are sequenced andcompared. In another embodiment, select regions such as multilocussequence tags (MLST), specific genes, or sets of genes may begenetically compared. In 16S embodiments, OTUs that share ≥97% averagenucleotide identity across the entire 16S or some variable region of the16S are considered the same OTU (see, e.g., Claesson et al. (2010)NUCLEIC ACIDS RES. 38: e200; Konstantinidis et al. (2006) PHILOS. TRANS.R. SOC. LOND. B BIOL. SCI. 361: 1929-1940). In embodiments involving thecomplete genome, MLSTs, specific genes, or sets of genes OTUs that share≥95% average nucleotide identity are considered the same OTU (see, e.g.,Achtman and Wagner (2008) NAT. REV. MICROBIOL. 6: 431-440;Konstantinidis et al. (2006)). OTUs are frequently defined by comparingsequences between organisms. Generally, sequences with less than 95%sequence identity are not considered to form part of the same OTU. OTUsmay also be characterized by any combination of nucleotide markers orgenes, in particular highly conserved genes (e.g., “house-keeping”genes), or a combination thereof. Such characterization employs, e.g.,WGS data or a whole genome sequence.

The term “phylogenetic diversity” refers to the biodiversity present ina given network ecology, core network ecology or network class ecologybased on the OTUs that comprise the network. Phylogenetic diversity is arelative term, meaning that a network ecology, core network or networkclass that is comparatively more phylogenetically diverse than anothernetwork contains a greater number of unique species, genera, andtaxonomic families. Uniqueness of a species, genera, or taxonomic familyis generally defined using a phylogenetic tree that represents thegenetic diversity all species, genera, or taxonomic families relative toone another. In another embodiment phylogenetic diversity may bemeasured using the total branch length or average branch length of aphylogenetic tree.

Phylogenetic diversity may be optimized in a bacterial composition byincluding a wide range of biodiversity.

“Phylogenetic tree”, “rDNA”, “rRNA”, “16S-rDNA”, “16S-rRNA” “16S”, “16Ssequencing”, “16S-NGS”, “18S”, “18S-rRNA”, “18S-rDNA” “18S sequencing”,and “18S-NGS” refer to the nucleic acids that encode for the RNAsubunits of the ribosome. rDNA refers to the gene that encodes the rRNAthat comprises the RNA subunits. There are two RNA subunits in theribosome termed the small subunit (SSU) and large subunit (LSU); the RNAgenetic sequences (rRNA) of these subunits are related to the gene thatencodes them (rDNA) by the genetic code. rDNA genes and theircomplementary RNA sequences are widely used for determination of theevolutionary relationships amount organisms as they are variable, yetsufficiently conserved to allow cross organism molecular comparisons.

Typically 16S rDNA sequence (approximately 1542 nucleotides in length)of the 30S SSU is used for molecular-based taxonomic assignments ofprokaryotes and the 18S rDNA sequence (approximately 1869 nucleotides inlength) of 40S SSLU is used for eukaryotes. The bacterial 16S rDNA isused in reconstructing the evolutionary relationships and sequencesimilarity of one bacterial isolate to another using phylogeneticapproaches. 16S sequences are used for phylogenetic reconstruction asthey are in general highly conserved, but contain specific hypervariableregions that harbor sufficient nucleotide diversity to differentiategenera and species of most bacteria.

The “V1-V9 regions” of the 16S rRNA refers to the first through ninthhypervariable regions of the 16S rRNA gene that are used for genetictyping of bacterial samples. These regions in bacteria are defined bynucleotides 69-99, 137-242, 433-497, 576-682, 822-879, 986-1043,1117-1173, 1243-1294 and 1435-1465 respectively using numbering based onthe E. coli system of nomenclature (see, e.g., Brosius et al. (1978)PROC. NAT'L. ACAD. SCI. USA 75(10): 4801-4805). In some embodiments, atleast one of the V1, V2, V3, V4, V5, V6, V7, V8, and V9 regions are usedto characterize an OTU. In one embodiment, the V1, V2, and V3 regionsare used to characterize an OTU. In another embodiment, the V3, V4, andV5 regions are used to characterize an OTU. In another embodiment, theV4 region is used to characterize an OTU. A person of ordinary skill inthe art can identify the specific hypervariable regions of a candidate16S rRNA by comparing the candidate sequence in question to a referencesequence and identifying the hypervariable regions based on similarityto the reference hypervariable regions, or alternatively, one can employWhole Genome Shotgun (WGS) sequence characterization of microbes or amicrobial community.

“Residual habitat products” refers to material derived from the habitatfor microbiota within or on a human or animal. For example, microbiotalive in feces in the gastrointestinal tract, on the skin itself, insaliva, mucus of the respiratory tract, or secretions of thegenitourinary tract (i.e., biological matter associated with themicrobial community). Substantially free of residual habitat productsmeans that the bacterial composition no longer contains the biologicalmatter associated with the microbial environment on or in the human oranimal subject and is 100% free, 99% free, 98% free, 97% free, 96% free,95% free, 94% free, 93% free, 92% free, 91% free, 90% free, 85% free,80% free, 75% free, 70% free, 65% free, or 60% free of any contaminatingbiological matter associated with the microbial community. Residualhabitat products can include abiotic materials (including undigestedfood) or it can include unwanted microorganisms. Substantially free ofresidual habitat products may also mean that the bacterial compositioncontains no detectable cells from a human or animal and that onlymicrobial cells are detectable. In one embodiment, substantially free ofresidual habitat products may also mean that the bacterial compositioncontains no detectable viral (e.g., bacterial viruses (i.e., phage)),fungal, or mycoplasmal contaminants. In another embodiment, it meansthat fewer than 1×10⁻²%, 1×10⁻³%, 1×10⁻⁴%, 1×10⁻⁵%, 1×10⁻⁶%, 1×10⁻⁷%,1×10⁻⁸% of the viable cells in the bacterial composition are human oranimal, as compared to microbial cells. There are multiple ways toaccomplish this degree of purity, none of which are limiting. Forexample, contamination may be reduced by isolating desired constituentsthrough multiple steps of streaking to single colonies on solid mediauntil replicate (such as, but not limited to, two) streaks from serialsingle colonies have shown only a single colony morphology.Alternatively, reduction of contamination can be accomplished bymultiple rounds of serial dilutions to single desired cells (e.g., adilution of 10⁻⁸ or 10⁻⁹), such as through multiple 10-fold serialdilutions. This can further be confirmed by showing that multipleisolated colonies have similar cell shapes and Gram staining behavior.Other methods for confirming adequate purity include genetic analysis(e.g., PCR and DNA sequencing), serology and antigen analysis, enzymaticand metabolic analysis, and methods using instrumentation such as flowcytometry with reagents that distinguish desired constituents fromcontaminants.

The term “subject” refers to any organism or animal subject that is anobject of a method or material, including mammals, e.g., humans,laboratory animals (e.g., primates, rats, mice, rabbits), livestock(e.g., cows, sheep, goats, pigs, turkeys, and chickens), household pets(e.g., dogs, cats, and rodents), horses, kangaroos, and transgenicnon-human animals. The subject may be suffering from a dysbiosis,including, but not limited to, an infection due to a gastrointestinalpathogen or may be at risk of developing or transmitting to others aninfection due to a gastrointestinal pathogen. Synonyms used hereininclude “patient” and “animal.” In some embodiments, the subject or hostmay be suffering from a dysbiosis, that contributes to or causes acondition classified as graft-versus-host disease, Crohn's disease,Celiac disease, inflammatory bowel disease, ulcerative colitis, multiplesclerosis, systemic lupus erythematosus, Sjogren's syndrome, or type 1diabetes. In some embodiments, the host may be suffering from metabolicendotoxemia, altered metabolism of primary bile acids, immune systemactivation, or an imbalance or reduced production of short chain fattyacids including, for example, butyrate, propionate, acetate, and abranched chain fatty acid.

The term “phenotype” refers to a set of observable characteristics of anindividual entity. As example an individual subject may have a phenotypeof “health” or “disease”. Phenotypes describe the state of an entity andall entities within a phenotype share the same set of characteristicsthat describe the phenotype. The phenotype of an individual results inpart, or in whole, from the interaction of the entities genome and/ormicrobiome with the environment.

“Spore” or “endospore” refers to an entity, particularly a bacterialentity, which is in a dormant, non-vegetative and non-reproductivestage. Spores are generally resistant to environmental stress such asradiation, desiccation, enzymatic treatment, temperature variation,nutrient deprivation, and chemical disinfectants.

A “spore population” refers to a plurality of spores present in acomposition. Synonymous terms used herein include spore composition,spore preparation, ethanol-treated spore fraction and spore ecology. Aspore population may be purified from a fecal donation, e.g., viaethanol or heat treatment, or a density gradient separation, or anycombination of methods described herein to increase the purity, potencyand/or concentration of spores in a sample. Alternatively, a sporepopulation may be derived through culture methods starting from isolatedspore former species or spore former OTUs or from a mixture of suchspecies, either in vegetative or spore form.

A “sporulation induction agent” is a material or physical-chemicalprocess that is capable of inducing sporulation in a bacterium, eitherdirectly or indirectly, in a host organism and/or in vitro.

To “increase production of bacterial entities” includes an activity or asporulation induction agent. “Production” includes conversion ofvegetative bacterial cells into spores and augmentation of the rate ofsuch conversion, as well as decreasing the germination of bacteria inspore form, decreasing the rate of spore decay in vivo, or ex vivo, orto increasing the total output of spores (e.g., via an increase involumetric output of fecal material).

“Synergy” or “synergistic interactions” refers to the interaction orcooperation of two or more microbes to produce a combined effect greaterthan the sum of their separate effects. In one embodiment, “synergy”between two or more microbes can result in the inhibition of a pathogensability to grow.

As used herein the term “vitamin” is understood to include any ofvarious fat-soluble or water-soluble organic substances (non-limitingexamples include vitamin A, Vitamin B1 (thiamine), Vitamin B2(riboflavin), Vitamin B3 (niacin or niacinamide), Vitamin B5(pantothenic acid), Vitamin B6 (pyridoxine, pyridoxal, or pyridoxamine,or pyridoxine hydrochloride), Vitamin B7 (biotin), Vitamin B9 (folicacid), and Vitamin B12 (various cobalamins; commonly cyanocobalamin invitamin supplements), vitamin C, vitamin D, vitamin E, vitamin K, K1 andK2 (i.e., MK-4, MK-7), folic acid and biotin) essential in minuteamounts for normal growth and activity of the body and obtainednaturally from plant and animal foods, or synthetically made,pro-vitamins, derivatives, and/or analogs.

As used herein, the term “antioxidant” is understood to include any oneor more of various substances such as beta-carotene (a vitamin Aprecursor), vitamin C, vitamin E, and selenium) that inhibit oxidationor reactions promoted by reactive oxygen species (“ROS”) and otherradical and non-radical species. Additionally, antioxidants aremolecules capable of slowing or preventing the oxidation of othermolecules. Non-limiting examples of antioxidants include astaxanthin,carotenoids, coenzyme Q10 (“CoQ10”), flavonoids, glutathione, Goji(wolfberry), hesperidin, lactowolfberry, lignan, lutein, lycopene,polyphenols, selenium, vitamin A, vitamin C, vitamin E, zeaxanthin, orcombinations thereof.

“Graft versus host disease” as used herein is an immunological disorderin which the immune cells of a transplant attack the tissues of atransplant recipient and may lead to organ dysfunction.

“Acute GVHD” as used herein is GVHD that prevents within the first 100days of transplant.

“Chronic GVHD” as used herein is GVHD that prevents after the first 100days of transplant.

“Inhibit,” “inhibiting,” and “inhibition” mean to decrease an activity,response, condition, disease, or other biological parameter. This caninclude but is not limited to the complete ablation of the activity,response, condition, or disease. This may also include, for example, a10% reduction in the activity, response, condition, or disease ascompared to the native or control level. Thus, the reduction can be a10, 20, 30, 40, 50, 60, 70, 80, 90, 100%, or any amount of reduction inbetween as compared to native or control levels.

“Treatment”, “treat”, or “treating”, mean a method of reducing theeffects of a disease or condition. Treatment can also refer to a methodof reducing the disease or condition itself rather than just thesymptoms. The treatment can be any reduction from native levels and canbe but is not limited to the complete ablation of the disease,condition, or the symptoms of the disease or condition. Therefore, inthe disclosed methods, treatment” can refer to a 10%, 20%, 30%, 40%,50%, 60%, 70%, 80%, 90%, or 100% reduction in the severity of anestablished disease or the disease progression. For example, a disclosedmethod for reducing the effects of GVHD is considered to be a treatmentif there is a 10% reduction in one or more symptoms of the disease in asubject with GVHD when compared to native levels in the same subject orcontrol subjects. Thus, the reduction can be a 10, 20, 30, 40, 50, 60,70, 80, 90, 100%, or any amount of reduction in between, as compared tonative or control levels. It is understood and herein contemplated that“treatment” does not necessarily refer to a cure of the disease orcondition, but an improvement in the outlook of a disease or condition(e.g., GVHD).

As used herein “preventing” or “prevention” refers to any methodologywhere the disease state does not occur due to the actions of themethodology (such as, for example, administration of a pharmaceuticalcomposition as described herein). In one aspect, it is understood thatprevention can also mean that the disease is not established to theextent that occurs in untreated controls. For example, there can be a5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%reduction in the establishment of disease.

As used herein, the term “recipient” refers to the subject that receivesa bone marrow or a solid organ transplantation.

Pharmaceutical Compositions of the Invention

Disclosed herein are pharmaceutical compositions, e.g., probioticcompositions, comprising a population of bacterial cells, e.g., animmunomodulatory bacterial cell population, such as an anti-inflammatorybacterial population, with or without one or more prebiotics, for theprevention, control, and treatment of inflammation, autoimmune andinflammatory disorders, dysbiosis, e.g., gastrointestinal or distaldysbiosis, disorders associated with dysbiosis, and for generalnutritional health. These compositions are advantageous in beingsuitable for safe administration to humans and other mammalian subjectsand are efficacious for the treatment, prevention, reduction of onsetand amelioration of inflammation, autoimmune and inflammatory disorders,dysbiosis, e.g., gastrointestinal or distal dysbiosis, disordersassociated with dysbiosis, and for general nutritional health. Thesepharmaceutical compositions are formulated as provided herein, andadministered to mammalian subjects using the methods as provided herein.In some embodiments, the compositions described herein are formulatedfor oral administration. In other embodiments, the compositionsdescribed herein are formulated for rectal administration.

In one embodiment, therapeutic compositions (e.g., pharmaceuticalcompositions) are provided for the treatment, prevention, reduction ofonset, and amelioration of, inflammation or one or more symptom of anautoimmune or inflammatory disorder, dysbiosis, e.g., gastrointestinalor distal dysbiosis, or a disorder associated with dysbiosis. As usedherein, “therapeutic” compositions include compositions that function ina prophylactic (e.g., preventative) manner. Generally, the population isprovided in an amount effective to treat (including to prevent) adisease, disorder or condition associated with or characterized byinflammation, dysbiosis, e.g., gastrointestinal or distal dysbiosis,inflammation, or an autoimmune or inflammatory disorder. Such treatmentmay be effective to reduce the severity of at least one symptom of thedysbiosis, e.g., gastrointestinal or distal dysbiosis, or an autoimmuneor inflammatory disorder. Such treatment may be effective to modulatethe microbiota diversity present in the mammalian recipient.

In some embodiments, the population of anti-inflammatory bacterial cellsis a purified population of bacterial cells. In some embodiments, saidpurified population of bacterial cells is isolated from a mammaliansource. In some embodiments, said purified population of bacterial cellsis isolated from a human source. In some embodiments, said purifiedpopulation of bacterial cells is isolated from the skin of a humansource. In some embodiments, said purified population of bacterial cellsis isolated from the gastrointestinal tract of a human source. In someembodiments, said purified population of bacterial cells is isolatedfrom the fecal matter of a subject. In some embodiments, said purifiedpopulation of bacterial cells is isolated from human fecal matter. Inother embodiments, said purified population of bacterial cells is notisolated from human fecal matter. In some embodiments, said purifiedpopulation of bacterial cells is not derived from human fecal matter.

In embodiments, the pharmaceutical compositions (e.g., probioticcompositions) contain immunomodulatory bacterial cells (e.g.,anti-inflammatory bacterial cells), which are capable of altering theimmune activity of a mammalian subject. In exemplary embodiments, theimmunomodulatory bacterial cells are capable of reducing inflammation ina mammalian subject. Such immunomodulatory bacterial cells are referredto herein as “anti-inflammatory bacteria” or “anti-inflammatorybacterial cells”. Immunomodulatory bacterial cells can act to alter theimmune activity of a subject directly or indirectly. For example,immunomodulatory bacteria can act directly on immune cells throughreceptors for bacterial components (e.g. Toll-like receptors) or byproducing metabolites such as immunomodulatory short chain fatty acids(SCFAs). Such SCFAs can have many positive impacts on the health of thesubject, by, for example, reducing inflammation, or improving intestinalbarrier integrity. Immunomodulatory bacterial cells can also impact theimmune activity of a subject by producing glutathione orgamma-glutamylcysteine.

Pharmaceutical compositions (e.g., probiotic compositions) containingimmunomodulatory bacteria (i.e., bacterial cells) can additionally oralternatively impact the immune activity of a subject indirectly bymodulating the activity of immune cells in the subject. For example,immunomodulatory bacteria may alter cytokine expression by host immunecells (e.g., macrophages, B lymphocytes, T lymphocytes, mast cells,peripheral blood mononuclear cells (PBMCs), etc.) or other types of hostcells capable of cytokine secretion (e.g., endothelial cells,fibroblasts, stromal cells, etc.). In an exemplary embodiment,pharmaceutical compositions (e.g., probiotic compositions) contain apopulation of anti-inflammatory bacterial cells that are capable ofinducing secretion of a anti-inflammatory cytokine by host cells (e.g.,host immune cells). For example, anti-inflammatory bacterial cells caninduce secretion of one or more anti-inflammatory cytokines such as, butnot limited to, IL-10, IL-13, IL-9, IL-4, IL-5, TGFβ, and combinationsthereof, by host cells (e.g., host immune cells). In another exemplaryembodiment, pharmaceutical compositions (e.g., probiotic compositions)contain anti-inflammatory bacterial cells that are capable of reducingsecretion of one or more pro-inflammatory cytokines by a host cell(e.g., by a host immune cell). For example, anti-inflammatory bacterialcells can reduce secretion of one or more pro-inflammatory cytokines,such as, but not limited to, IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1,MIP1α, MIP1β, TNFα, and combinations thereof, by host cells (e.g., hostimmune cells). In some embodiments, the induction and/or secretion ofsaid pro-inflammatory cytokines may be induced by (e.g., in response to,either directly or indirectly) a bacteria (e.g., Enterococcus faecalis).Other cytokines that may be modulated by immunomodulatory bacterialcells include, for example, IL-17A, IL-2, and IL-9.

In some embodiments, immunomodulatory bacteria (i.e., immunomodulatorybacterial cells) are selected for inclusion in a pharmaceuticalcomposition (e.g., probiotic composition) of the invention based on thedesired effect of the immunomodulatory bacteria on cytokine secretion bya host cell or a population of host cells (e.g., a host immune cell(e.g., a PBMC)). In some embodiments, said effect of theimmunomodulatory bacteria is assessed in vitro using a population ofhost cells (e.g., a population of isolated host immune cells). Forexample, in one embodiment, a probiotic composition containsanti-inflammatory bacteria that increase secretion of aanti-inflammatory cytokine, for example, IL-10, IL-13, IL-9, IL-4, IL-5,TGFβ, and combinations thereof, by a host cell (e.g., a host immune cell(e.g., PBMCs, macrophages, B lymphocytes, T lymphocytes, mast cells). Insome embodiments, the anti-inflammatory bacteria increase secretion oftwo or more anti-inflammatory cytokines. In some embodiments, theanti-inflammatory bacteria increase secretion of three or moreanti-inflammatory cytokines. In some embodiments, the anti-inflammatorybacteria increase secretion of four or more anti-inflammatory cytokines.In some embodiments, the anti-inflammatory bacteria increase secretionof five or more anti-inflammatory cytokines. In exemplary embodiments,the increase is an increase of at least 5%, 10%, 20%, 30%, 40%, 50%,60%, 70%, 80%, 80%, 100%, 200%, 300%, 500% or more. In otherembodiments, a pharmaceutical composition contains anti-inflammatorybacteria that decrease secretion of a pro-inflammatory cytokine, forexample, IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1, MIP1α, MIP1β, TNFα,and combinations thereof, by a host cell. In some embodiments, theanti-inflammatory bacteria decrease secretion of five or morepro-inflammatory cytokines. In exemplary embodiments, the decrease is adecrease of at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 80%,100%, 200%, 300%, 500% or more. In another embodiment, thepharmaceutical composition contains anti-inflammatory bacterial cellsthat increase secretion of one or more anti-inflammatory cytokines andreduce secretion of one or more pro-inflammatory cytokines by a hostcell (e.g., a host immune cell). Alterations in cytokine expression mayoccur locally, e.g., in the gastrointestinal tract of a subject, or at asite distal to a microbial niche, e.g., distal to the gastrointestinaltract. In some embodiments, the induction and/or secretion of saidpro-inflammatory cytokines may be induced by (e.g., in response to,either directly or indirectly) a bacteria (e.g., Enterococcus faecalis).

In some aspects, the pharmaceutical compositions described herein and/ora prebiotic (e.g., a carbohydrate) modulate the release of immunestimulatory cytokines by host cells (e.g., host immune cells). Inpreferred embodiments, the administered immunomodulatory bacterial cells(e.g., anti-inflammatory bacterial cells) and/or a prebiotic (e.g., acarbohydrate) inhibit or reduce the release of immune stimulatorycytokines. Non-limiting examples of immune modulating cytokines andligands include B lymphocyte chemoattractant (“BLC”), C-C motifchemokine 11 (“Eotaxin-1”), Eosinophil chemotactic protein 2(“Eotaxin-2”), Granulocyte colony-stimulating factor (“G-CSF”),Granulocyte macrophage colony-stimulating factor (“GM-CSF”), 1-309,Intercellular Adhesion Molecule 1 (“ICAM-1”), Interferon gamma(“IFN-γ”), Interlukin-1 alpha (“IL-1α”), Interlukin-1 (“IL-1”),Interleukin 1 receptor antagonist (“IL-1 ra”), Interleukin-2 (“IL-2”),Interleukin-4 (“IL-4”), Interleukin-5 (“IL-5”), Interleukin-6 (“IL-6”),Interleukin-6 soluble receptor (“IL-6 sR”), Interleukin-7 (“IL-7”),Interleukin-8 (“IL-8”), Interleukin-10 (“IL-10”), Interleukin-11(“IL-11”), Subunit R of Interleukin-12 (“IL-12 p40” or “IL-12 p70”),Interleukin-13 (“IL-13”), Interleukin-15 (“IL-15”), Interleukin-16(“IL-16”), Interleukin-17 (“IL-17”), Chemokine (C-C motif) Ligand 2(“MCP-1”), Macrophage colony-stimulating factor (“M-CSF”), Monokineinduced by gamma interferon (“MIG”), Chemokine (C-C motif) ligand 2(“MIP-1 alpha”), Chemokine (C-C motif) ligand 4 (“MIP-10”), Macrophageinflammatory protein-1-6 (“MIP-16”), Platelet-derived growth factorsubunit B (“PDGF-BB”), Chemokine (C-C motif) ligand 5, Regulated onActivation, Normal T cell Expressed and Secreted (“RANTES”), TIMPmetallopeptidase inhibitor 1 (“TIMP-1”), TIMP metallopeptidase inhibitor2 (“TIMP-2”), Tumor necrosis factor, lymphotoxin-α (“TNF-α”), Tumornecrosis factor, lymphotoxin-0 (“TNF R”), Soluble TNF receptor type 1(“sTNFRI”), sTNFRIIAR, Brain-derived neurotrophic factor (“BDNF”), Basicfibroblast growth factor (“bFGF”), Bone morphogenetic protein 4(“BMP-4”), Bone morphogenetic protein 5 (“BMP-5”), Bone morphogeneticprotein 7 (“BMP-7”), Nerve growth factor (“b-NGF”), Epidermal growthfactor (“EGF”), Epidermal growth factor receptor (“EGFR”),Endocrine-gland-derived vascular endothelial growth factor (“EG-VEGF”),Fibroblast growth factor 4 (“FGF-4”), Keratinocyte growth factor(“FGF-7”), Growth differentiation factor 15 (“GDF-15”), Glialcell-derived neurotrophic factor (“GDNF”), Growth Hormone,Heparin-binding EGF-like growth factor (“HB-EGF”), Hepatocyte growthfactor (“HGF”), Insulin-like growth factor binding protein 1(“IGFBP-1”), Insulin-like growth factor binding protein 2 (“IGFBP-2”),Insulin-like growth factor binding protein 3 (“IGFBP-3”), Insulin-likegrowth factor binding protein 4 (“IGFBP-4”), Insulin-like growth factorbinding protein 6 (“IGFBP-6”), Insulin-like growth factor 1 (“IGF-1”),Insulin, Macrophage colony-stimulating factor (“M-CSF R”), Nerve growthfactor receptor (“NGF R”), Neurotrophin-3 (“NT-3”), Neurotrophin-4(“NT-4”), Osteoclastogenesis inhibitory factor (“Osteoprotegerin”),Platelet-derived growth factor receptors (“PDGF-AA”),Phosphatidylinositol-glycan biosynthesis (“PIGF”), Skp, Cullin, F-boxcontaining comples (“SCF”), Stem cell factor receptor (“SCF R”),Transforming growth factor α (“TGF-α”), Transforming growth factor β-1(“TGF β1”), Transforming growth factor β-3 (“TGF β3”), Vascularendothelial growth factor (“VEGF”), Vascular endothelial growth factorreceptor 2 (“VEGFR2”), Vascular endothelial growth factor receptor 3(“VEGFR3”), VEGF-D 6Ckine, Tyrosine-protein kinase receptor UFO (“Axl”),Betacellulin (“BTC”), Mucosae-associated epithelial chemokine (“CCL28”),Chemokine (C-C motif) ligand 27 (“CTACK”), Chemokine (C—X-C motif)ligand 16 (“CXCL16”), C-X-C motif chemokine 5 (“ENA-78”), Chemokine (C-Cmotif) ligand 26 (“Eotaxin-3”), Granulocyte chemotactic protein 2(“GCP-2”), GRO, Chemokine (C-C motif) ligand 14 (“HCC-1”), Chemokine(C-C motif) ligand 16 (“HCC-4”), Interleukin-9 (“IL-9”), Interleukin-17F (“IL-17F”), Interleukin-18-binding protein (“IL-18 BPa”),Interleukin-28 A (“IL-28A”), Interleukin 29 (“IL-29”), Interleukin 31(“IL-31”), C-X-C motif chemokine 10 (“IP-10”), Chemokine receptor CXCR3(“I-TAC”), Leukemia inhibitory factor (“LIF”), Light, Chemokine (Cmotif) ligand (“Lymphotactin”), Monocyte chemoattractant protein 2(“MCP-2”), Monocyte chemoattractant protein 3 (“MCP-3”), Monocytechemoattractant protein 4 (“MCP-4”), Macrophage-derived chemokine(“MDC”), Macrophage migration inhibitory factor (“MIF”), Chemokine (C-Cmotif) ligand 20 (“MIP-3 α”), C-C motif chemokine 19 (“MIP-3 β”),Chemokine (C-C motif) ligand 23 (“MPIF-1”), Macrophage stimulatingprotein alpha chain (“MSP-α”), Nucleosome assembly protein 1-like 4(“NAP-2”), Secreted phosphoprotein 1 (“Osteopontin”), Pulmonary andactivation-regulated cytokine (“PARC”), Platelet factor 4 (“PF4”),Stroma cell-derived factor-1 α (“SDF-1 α”), Chemokine (C-C motif) ligand17 (“TARC”), Thymus-expressed chemokine (“TECK”), Thymic stromallymphopoietin (“TSLP 4-IBB”), CD 166 antigen (“ALCAM”), Cluster ofDifferentiation 80 (“B7-1”), Tumor necrosis factor receptor superfamilymember 17 (“BCMA”), Cluster of Differentiation 14 (“CD14”), Cluster ofDifferentiation 30 (“CD30”), Cluster of Differentiation 40 (“CD40Ligand”), Carcinoembryonic antigen-related cell adhesion molecule 1(biliary glycoprotein) (“CEACAM-1”), Death Receptor 6 (“DR6”),Deoxythymidine kinase (“Dtk”), Type 1 membrane glycoprotein(“Endoglin”), Receptor tyrosine-protein kinase erbB-3 (“ErbB3”),Endothelial-leukocyte adhesion molecule 1 (“E-Selectin”), Apoptosisantigen 1 (“Fas”), Fms-like tyrosine kinase 3 (“Flt-3L”), Tumor necrosisfactor receptor superfamily member 1 (“GITR”), Tumor necrosis factorreceptor superfamily member 14 (“HVEM”), Intercellular adhesion molecule3 (“ICAM-3”), IL-1 R4, IL-1 RI, IL-10 Rβ, IL-17R, IL-2Rγ, IL-21R,Lysosome membrane protein 2 (“LIMPII”), Neutrophil gelatinase-associatedlipocalin (“Lipocalin-2”), CD62L (“L-Selectin”), Lymphatic endothelium(“LYVE-1”), MHC class I polypeptide-related sequence A (“MICA”), MHCclass I polypeptide-related sequence B (“MICB”), NRGl-β1, Beta-typeplatelet-derived growth factor receptor (“PDGF Rβ”), Plateletendothelial cell adhesion molecule (“PECAM-1”), RAGE, Hepatitis A viruscellular receptor 1 (“TIM-1”), Tumor necrosis factor receptorsuperfamily member IOC (“TRAIL R3”), Trappin protein transglutaminasebinding domain (“Trappin-2”), Urokinase receptor (“uPAR”), Vascular celladhesion protein 1 (“VCAM-1”), XEDARActivin A, Agouti-related protein(“AgRP”), Ribonuclease 5 (“Angiogenin”), Angiopoietin 1, Angiostatin,Catheprin S, CD40, Cryptic family protein IB (“Cripto-1”), DAN,Dickkopf-related protein 1 (“DKK-1”), E-Cadherin, Epithelial celladhesion molecule (“EpCAM”), Fas Ligand (FasL or CD95L), Fcg RIIB/C,FoUistatin, Galectin-7, Intercellular adhesion molecule 2 (“ICAM-2”),IL-13 R1, IL-13R2, IL-17B, IL-2 Ra, IL-2 Rb, IL-23, LAP, Neuronal celladhesion molecule (“NrCAM”), Plasminogen activator inhibitor-1(“PAI-1”), Platelet derived growth factor receptors (“PDGF-AB”),Resistin, stromal cell-derived factor 1 (“SDF-1 β”), sgpl30, Secretedfrizzled-related protein 2 (“ShhN”), Sialic acid-bindingimmunoglobulin-type lectins (“Siglec-5”), ST2, Transforming growthfactor-β 2 (“TGF β 2”), Tie-2, Thrombopoietin (“TPO”), Tumor necrosisfactor receptor superfamily member 10D (“TRAIL R4”), Triggering receptorexpressed on myeloid cells 1 (“TREM-1”), Vascular endothelial growthfactor C (“VEGF-C”), VEGFRlAdiponectin, Adipsin (“AND”), α-fetoprotein(“AFP”), Angiopoietin-like 4 (“ANGPTL4”), β-2-microglobulin (“B2M”),Basal cell adhesion molecule (“BCAM”), Carbohydrate antigen 125(“CA125”), Cancer Antigen 15-3 (“CA15-3”), Carcinoembryonic antigen(“CEA”), cAMP receptor protein (“CRP”), Human Epidermal Growth FactorReceptor 2 (“ErbB2”), Follistatin, Follicle-stimulating hormone (“FSH”),Chemokine (C-X-C motif) ligand 1 (“GRO a”), human chorionic gonadotropin(“3 HCG”), Insulin-like growth factor 1 receptor (“IGF-1 sR”), IL-1sRII, IL-3, IL-18 Rb, IL-21, Leptin, Matrix metalloproteinase-1(“MMP-1”), Matrix metalloproteinase-2 (“MMP-2”), Matrixmetalloproteinase-3 (“MMP-3”), Matrix metalloproteinase-8 (“MMP-8”),Matrix metalloproteinase-9 (“MMP-9”), Matrix metalloproteinase-10(“MMP-10”), Matrix metalloproteinase-13 (“MMP-13”), Neural Cell AdhesionMolecule (“NCAM-1”), Entactin (“Nidogen-1”), Neuron specific enolase(“NSE”), Oncostatin M (“OSM”), Procalcitonin, Prolactin, Prostatespecific antigen (“PSA”), Sialic acid-binding Ig-like lectin 9(“Siglec-9”), ADAM 17 endopeptidase (“TACE”), Thyroglobulin,Metalloproteinase inhibitor 4 (“TIMP-4”), TSH2B4, Disintegrin andmetalloproteinase domain-containing protein 9 (“ADAM-9”), Angiopoietin2, Tumor necrosis factor ligand superfamily member 13/Acidicleucine-rich nuclear phosphoprotein 32 family member B (“APRIL”), Bonemorphogenetic protein 2 (“BMP-2”), Bone morphogenetic protein 9(“BMP-9”), Complement component 5a (“C5a”), Cathepsin L, CD200, CD97,Chemerin, Tumor necrosis factor receptor superfamily member 6B (“DcR3”),Fatty acid-binding protein 2 (“FABP2”), Fibroblast activation protein,alpha (“FAP”), Fibroblast growth factor 19 (“FGF-19”), Galectin-3,Hepatocyte growth factor receptor (“HGF R”), IFN-γα/β R2, Insulin-likegrowth factor 2 (“IGF-2”), Insulin-like growth factor 2 receptor (“IGF-2R”), Interleukin-1 receptor 6 (“IL-1R6”), Interleukin 24 (“IL-24”),Interleukin 33 (“IL-33”, Kallikrein 14, Asparaginyl endopeptidase(“Legumain”), Oxidized low-density lipoprotein receptor 1 (“LOX-1”),Mannose-binding lectin (“MBL”), Neprilysin (“NEP”), Notch homolog 1,translocation-associated (Drosophila) (“Notch-1”), Nephroblastomaoverexpressed (“NOV”), Osteoactivin, Programmed cell death protein 1(“PD-1”), N-acetylmuramoyl-L-alanine amidase (“PGRP-5”), Serpin A4,Secreted frizzled related protein 3 (“sFRP-3”), Thrombomodulin,Toll-like receptor 2 (“TLR2”), Tumor necrosis factor receptorsuperfamily member 10A (“TRAIL Rl”), Transferrin (“TRF”), WIF-lACE-2,Albumin, AMICA, Angiopoietin 4, B-cell activating factor (“BAFF”),Carbohydrate antigen 19-9 (“CA19-9”), CD 163, Clusterin, CRT AM,Chemokine (C-X-C motif) ligand 14 (“CXCL14”), Cystatin C, Decorin(“DCN”), Dickkopf-related protein 3 (“Dkk-3”), Delta-like protein 1(“DLL1”), Fetuin A, Heparin-binding growth factor 1 (“aFGF”), Folatereceptor α (“FOLR1”), Furin, GPCR-associated sorting protein 1(“GASP-1”), GPCR-associated sorting protein 2 (“GASP-2”), Granulocytecolony-stimulating factor receptor (“GCSF R”), Serine protease hepsin(“HAI-2”), Interleukin-17B Receptor (“IL-17B R”), Interleukin 27(“IL-27”), Lymphocyte-activation gene 3 (“LAG-3”), Apolipoprotein A-V(“LDL R”), Pepsinogen I, Retinol binding protein 4 (“RBP4”), SOST,Heparan sulfate proteoglycan (“Syndecan-1”), Tumor necrosis factorreceptor superfamily member 13B (“TACI”), Tissue factor pathwayinhibitor (“TFPI”), TSP-1, Tumor necrosis factor receptor superfamily,member 10b (“TRAIL R2”), TRANCE, Troponin I, Urokinase PlasminogenActivator (“uPA”), Cadherin 5, type 2 or VE-cadherin (vascularendothelial) also known as CD144 (“VE-Cadherin”),WNTl-inducible-signaling pathway protein 1 (“WISP-1”), and ReceptorActivator of Nuclear Factor κ B (“RANK”).

In other embodiments, pharmaceutical compositions (e.g., probioticcompositions) containing immunomodulatory bacteria that impact theimmune activity of a subject by promoting the differentiation and/orexpansion of particular subpopulations of immune cells. For example,immunomodulatory bacteria can increase or decrease the proportion ofTreg cells, Th17 cells, Th1 cells, or Th2 cells in a subject. Theincrease or decrease in the proportion of immune cell subpopulations maybe systemic, or it may be localized to a site of action of theprobiotic, e.g., in the gastrointestinal tract or at a site of distaldysbiosis. In some embodiments, immunomodulatory bacteria (i.e.,anti-inflammatory bacterial cells) are selected for inclusion in apharmaceutical composition (e.g., a probiotic composition) of theinvention based on the desired effect of the pharmaceutical compositionon the differentiation and/or expansion of subpopulations of immunecells in the subject.

In one embodiment, a pharmaceutical composition (e.g., a probioticcomposition) contains immunomodulatory bacteria (i.e., immunomodulatorybacterial cells) that increase the proportion of Treg cells in a subject(e.g., by inducing expansion of Treg cells in the subject). In anotherembodiment, a pharmaceutical composition contains immunomodulatorybacteria that decrease the proportion of Treg cells in a subject. In oneembodiment, a pharmaceutical composition contains immunomodulatorybacteria that increase the proportion of Th17 cells in a subject (e.g.,by inducing expansion of Th17 cells in the subject). In anotherembodiment, a pharmaceutical composition contains immunomodulatorybacteria that decrease the proportion of Th17 cells in a subject. In oneembodiment, a pharmaceutical composition contains immunomodulatorybacteria that increase the proportion of Th1 cells in a subject (e.g.,by inducing expansion of Th1 cells in the subject). In anotherembodiment, a pharmaceutical composition contains immunomodulatorybacteria that decrease the proportion of Th1 cells in a subject. In oneembodiment, a pharmaceutical composition contains immunomodulatorybacteria that increase the proportion of Th2 cells in a subject (e.g.,by inducing expansion of Th2 cells in the subject). In anotherembodiment, a pharmaceutical composition contains immunomodulatorybacteria that decrease the proportion of Th2 cells in a subject. Theincrease or decrease in the proportion of immune cell subpopulations maybe systemic, or it may be localized to a site of action of theprobiotic, e.g., in the gastrointestinal tract or at a site of distaldysbiosis.

In one embodiment, a pharmaceutical composition (e.g., a probioticcomposition) contains immunomodulatory bacteria capable of modulatingthe proportion of one or more populations of Treg cells, Th17 cells, Th1cells, Th2 cells, and combinations thereof, in a subject. Certain immunecell profiles may be particularly desirable to treat or preventparticular disorders associated with a dysbiosis. For example, treatmentor prevention of an autoimmune or inflammatory disorder (e.g., GVHD) canbe promoted by increasing the quantity of Treg cells and Th2 cells, anddecreasing the quantity of Th17 cells and Th1 cells. Accordingly,pharmaceutical compositions (e.g., probiotic compositions) for thetreatment or prevention of an autoimmune or inflammatory disorder (e.g.,GVHD) contain immunomodulatory bacteria capable of promoting thedifferentiation and/or expansion of Treg cells and Th2 cells, andreducing Th17 and Th1 cells in the subject.

In one embodiment, pharmaceutical compositions (e.g., a therapeuticprobiotic compositions) containing a purified population ofimmunomodulatory microbes, e.g., immunomodulatory bacterial cells, areprovided, with or without one or more prebiotics, in an amount effectiveto: i) treat or prevent dysbiosis, e.g., gastrointestinal or distaldysbiosis, inflammation, or an autoimmune or inflammatory disorder;and/or ii) augment at least one type of microbe, e.g., a bacterium, notpresent in the therapeutic composition in a mammalian recipient subjectto whom the pharmaceutical composition is administered; and/or iii)engraft at least one type of microbe, e.g., an anti-inflammatorybacterial cell, present in the therapeutic composition but not presentin a mammalian subject prior to treatment.

In another embodiment, pharmaceutical compositions containing a purifiedpopulation of immunomodulatory bacteria (e.g., anti-inflammatorybacterial cells) are provided, in an amount effective to i) augment themicrobiota diversity present in the mammalian recipient and/or ii) treator prevent dysbiosis, e.g., gastrointestinal or distal dysbiosis,inflammation, or an autoimmune or inflammatory disorder in a mammalianrecipient subject to whom the therapeutic composition is administered,wherein the purified population of immunomodulatory bacteria is obtainedby separating the population from at least one residual habitat productin a fecal material obtained from one or a plurality of mammalian donorsubjects. In some embodiments, individual bacterial strains can becultured from fecal material. These strains can then be purified orotherwise isolated and used singly or in combination. In one embodiment,the probiotic composition does not contain a fecal extract.

In one embodiment, the pharmaceutical compositions described herein maybe used to treat or correct a dysbiosis in a subject. The dysbiosis maybe, for example, a local dysbiosis, or a distal dysbiosis. In anotherembodiment, the probiotic compositions described herein may be used toprevent a dysbiosis in a subject at risk for developing a dysbiosis.

In another embodiment, pharmaceutical compositions containing a purifiedpopulation of immunomodulatory bacteria (e.g., anti-inflammatorybacterial cells) are provided, in an amount effective to i) augment themicrobiota diversity present in the mammalian recipient and/or ii) treator prevent dysbiosis, e.g., gastrointestinal or distal dysbiosis,inflammation, or an autoimmune or inflammatory disorder in a mammalianrecipient subject to whom the therapeutic composition is administered,wherein the purified population of immunomodulatory bacteria is obtainedby separating the population from a non-fecal material source.

In some embodiments, a pharmaceutical composition containing a purifiedpopulation of immunomodulatory bacterial cells (e.g., anti-inflammatorybacterial cells) described above is co-administered or co-formulatedwith one or more prebiotics, e.g., carbohydrates. In some embodiments, apharmaceutical composition is administered before one or more prebioticsis administered to a subject. In some embodiments, the pharmaceuticalcomposition is administered after one or more prebiotics is administeredto a subject. In some embodiments, a pharmaceutical compositioncontaining a purified population of immunomodulatory bacterial cells isadministered concurrently with one or more prebiotics. In otherembodiments, a pharmaceutical composition containing a purifiedpopulation of immunomodulatory bacterial cells is administeredsequentially with one or more prebiotics. In some embodiments, apurified population of immunomodulatory bacterial cells is administeredin a pharmaceutical composition formulated to contain one or morepharmaceutical excipients, and optionally one or more prebiotics.

Immunomodulatory bacterial cells (e.g., anti-inflammatory bacterialcells) involved in modulation of the host immune system i) may be humancommensals; ii) may be part of an organ's healthy-state microbiome; ii)may be part of a distal organ's healthy-state microbiome; iv) may beexogenous microbes; v) may be innocuous; vi) may be pathobionts; vii)may be pathogens; viii) may be opportunistic pathogens; or ix) anycombination thereof. In some aspects, microbes are not required to beactively proliferating (e.g., spores, dormant cells, cells with reducedmetabolic rate, or heat-killed cells) to have an immunomodulatoryeffect. In certain aspects, microbial cell components, rather than wholemicrobial cells, may have immunomodulatory effects. Non-limitingexamples of microbial components are lipids, carbohydrates, proteins,nucleic acids, and small molecules.

The pharmaceutical compositions provided herein, may optionally furthercomprise a prebiotic, a non-microbial immunomodulatory carbohydrates, ora microbial immunomodulatory cell component, that are effective for theprevention or treatment of an autoimmune or inflammatory disorder suchas graft-versus-host disease (GVHD), an inflammatory bowel disease (IBD)including, but not limited to, ulterative colitis and Crohn's disease,multiple sclerosis (MS), systemic lupus erythematosus (SLE), type Idiabetes, rheumatoid arthritis, Sjögren's syndrome, and Celiac disease,or dysbiosis.

In certain embodiments, the pharmaceutical compositions comprise atleast one type of immunomodulatory bacterial cells (e.g., at least onetype of anti-inflammatory bacterial cell) and, optionally, at least oneprebiotic (e.g., a carbohydrate), and optionally further comprise amicrobial immunomodulatory cell component or substrate for theproduction of immunomodulatory metabolites, that are effective for theprevention or treatment of an autoimmune or inflammatory disorder.Methods for the prevention and/or treatment of autoimmune andinflammatory diseases in human subjects are also disclosed herein.

In some embodiments, the pharmaceutical compositions, e.g., probioticcompositions, of the invention comprise purified spore populations ofanti-inflammatory bacterial cells. In one embodiment, the purified sporepopulations can engraft in the host and remain present for 1 day, 2days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 14 days, 21 days,25 days, 30 days, 60 days, 90 days, or longer than 90 days.Additionally, the purified spore populations can induce other healthycommensal bacteria found in a healthy gut to engraft in the host thatare not present in the purified spore populations or present at lesserlevels. Therefore, these species are considered to “augment” thedelivered spore populations. In this manner, commensal speciesaugmentation of the purified spore population in the recipient's gutleads to a more diverse population of gut microbiota than presentinitially.

Preferred bacterial cells for use in the present invention includebacterial cells of the genera Acetanaerobacterium, Acetivibrio,Alicyclobacillus, Alkaliphilus, Anaerofustis, Anaerosporobacter,Anaerostipes, Anaerotruncus, Anoxybacillus, Bacillus, Bacteroides,Blautia, Brachyspira, Brevibacillus, Bryantella, Bulleidia,Butyricicoccus, Butyrivibrio, Catenibacterium, Chlamydiales,Clostridiaceae, Clostridiales, Clostridium, Collinsella, Coprobacillus,Coprococcus, Coxiella, Deferribacteres, Desulfitobacterium,Desulfotomaculum, Dorea, Eggerthella, Erysipelothrix,Erysipelotrichaceae, Ethanoligenens, Eubacterium, Faecalibacterium,Filifactor, Flavonifractor, Flexistipes, Fulvimonas, Fusobacterium,Gemmiger, Geobacillus, Gloeobacter, Holdemania,Hydrogenoanaerobacterium, Kocuria, Lachnobacterium, Lachnospira,Lachnospiraceae, Lactobacillus, Lactonifactor, Leptospira, Lutispora,Lysinibacillus, Mollicutes, Moorella, Nocardia, Oscillibacter,Oscillospira, Paenibacillus, Papillibacter, Pseudoflavonifractor,Robinsoniella, Roseburia, Ruminococcaceae, Ruminococcus,Saccharomonospora, Sarcina, Solobacterium, Sporobacter,Sporolactobacillus, Streptomyces, Subdoligranulum, Sutterella,Syntrophococcus, Thermoanaerobacter, Thermobifida, and Turicibacter.

Preferred bacterial cells also include bacterial cells of the generaAcetonema, Alkaliphilus, Amphibacillus, Ammonifex, Anaerobacter,Caldicellulosiruptor, Caloramator, Candidatus, Carboxydibrachium,Carboxydothermus, Cohnella, Dendrosporobacter Desulfitobacterium,Desulfosporosinus, Halobacteroides, Heliobacterium, Heliophilum,Heliorestis, Lachnoanaerobaculum, Lysinibacillus, Oceanobacillus, Orenia(S.), Oxalophagus, Oxobacter, Pelospora, Pelotomaculum, Propionispora,Sporohalobacter, Sporomusa, Sporosarcina, Sporotomaculum,Symbiobacterium, Syntrophobotulus, Syntrophospora, Terribacillus,Thermoanaerobacter, and Thermosinus.

In another embodiment, a probiotic composition of the invention consistsessentially of Blautia.

In one embodiment, a probiotic composition of the invention does notcomprise Blautia alone.

As provided herein, the pharmaceutical compositions comprise, or in thealternative, modulate, the colonization and/or engraftment, of thefollowing exemplary bacterial entities (e.g., bacterial cells belongingto particular bacterial strains, bacterial species, or bacterialgenera): Lactobacillus gasseri, Lactobacillus fermentum, Lactobacillusreuteri, Enterococcus faecalis, Enterococcus durans, Enterococcusvillorum, Blautia luti, Blautia coccoides, Blautia hydrogenotrophica,Blautia hansenii, Blautia wexlerae, Lactobacillus plantarum, Pediococcusacidilactici, Staphylococcus pasteuri, Staphylococcus cohnii,Streptococcus sanguinis, Streptococcus sinensis, Streptococcus mitis,Streptococcus sp. SCA22, Streptococcus sp. CR-3145, Streptococcusanginosus, Streptococcus mutans, Coprobacillus cateniformis, Clostridiumsaccharogumia, Eubacterium dolichum DSM 3991, Clostridium sp. PPf35E6,Clostridium sordelli ATCC 9714, Ruminococcus torques, Ruminococcusgnavus, Clostridium clostridioforme, Ruminococcus obeum, Blautiaproducta, Clostridium sp. ID5, Megasphaera micronuciformis, Veillonellaparvula, Clostridium methylpentosum, Clostridium islandicum,Faecalibacterium prausnitzii, Bacteroides unformmis, Eubacteriumrectale, Bacteroides thetaiotaomicron, Bacteroides acidifaciens,Bacteroides ovatus, Bacteroides fragilis, Parabacteroides distasonis,Propinionibacteirum propionicum, Actinomycs hyovaginalis, Rothiamucilaginosa, Rothia aeria, Bifidobacterium breve, Scardovia inopinataand Eggerthella lenta.

Preferred bacterial strains are provided in Table 1, Table 1A, Table 1B,Table 1C, Table 1D, Table 1E, and Table 1F. Optionally, in someembodiments, preferred bacterial species are spore formers. Thebacterial cells may be in the vegetative form and/or in the spore form.Thus, in some embodiments, the bacterial cell is present in thepharmaceutical composition solely in spore form. In other embodiments,the bacterial cell is present in the pharmaceutical composition solelyin vegetative form. Yet, in other embodiments, the bacterial cell may bepresent in the pharmaceutical composition in a combination of vegetativeform and spore form. Where specific strains of a species are provided,one of skill in the art will recognize that other strains of the speciescan be substituted for the named strain.

In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Acidaminococcusintestine. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isAcinetobacter baumannii. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Acinetobacter lwoffii. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Akkermansia muciniphila. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Alistipes putredinis. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Alistipes shahii. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Anaerostipes hadrus. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Anaerotruncus colihominis.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Bacteroides caccae.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Bacteroidescellulosilyticus. In one embodiment, the bacterial entity, e.g., speciesor strain, useful in the compositions and methods of the invention isBacteroides dorei. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Bacteroides eggerthii. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Bacteroides finegoldii. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Bacteroides fragilis. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Bacteroides massiliensis.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Bacteroides ovatus.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Bacteroidessalanitronis. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isBacteroides salyersiae. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Bacteroides sp. 1_1_6. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Bacteroides sp. 3_1_23. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Bacteroides sp. D20. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Bacteroidesthetaiotaomicrond. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Bacteroides uniformis. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Bacteroides vulgatus. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Bifidobacterium adolescentis. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Bifidobacterium bifidum. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Bifidobacterium breve.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Bifidobacteriumfaecale. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isBifidobacterium kashiwanohense. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Bifidobacterium longum subsp. Longum. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Bifidobacteriumpseudocatenulatum. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Bifidobacterium stercoris. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Blautia (Ruminococcus) coccoides. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia faecis. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia glucerasea. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia (Ruminococcus)hansenii. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isBlautia hydrogenotrophica (Ruminococcus hydrogenotrophicus). In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia (Ruminococcus)luti. In one embodiment, the bacterial entity, e.g., species or strain,useful in the compositions and methods of the invention is Blautia(Ruminococcus) obeum. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Blautia producta (Ruminococcus productus). In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia (Ruminococcus)schinkii. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isBlautia stercoris. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Blautia uncultured bacterium clone BKLE_a03_2 (GenBank:EU469501.1). In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isBlautia uncultured bacterium clone SJTU_B_14_30 (GenBank: EF402926.1).In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Blautia unculturedbacterium clone SJTU_C_14_16 (GenBank: EF404657.1). In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia unculturedbacterium clone S1-5 (GenBank: GQ898099.1). In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Blautia uncultured PACO000178_s(www.ezbiocloud.net/eztaxon/hierarchy?m=browse&k=PAC000178&d=2). In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Blautia wexlerae. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Candidatus Arthromitus sp.SFB-mouse-Yit. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isCatenibacterium mitsuokai. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Clostridiaceae bacterium (Dielma fastidiosa) JC13. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Clostridiales bacterium1_7_47FAA. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isClostridium asparagiforme. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Clostridium bolteae. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Clostridium clostridioforme. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Clostridium glycyrrhizinilyticum. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Clostridium (Hungatella)hathewayi. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isClostridium histolyticum. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Clostridium indolis. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Clostridium leptum. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Clostridium (Tyzzerella) nexile. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Clostridium perfringens. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Clostridium(Erysipelatoclostridium) ramosum. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Clostridium scindens. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Clostridium septum. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Clostridium sp. 14774. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Clostridium sp.7_3_54FAA. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isClostridium sp. HGF2. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Clostridium symbiosum. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Collinsella aerofaciens. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Collinsella intestinalis. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Coprobacillus sp. D7. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Coprococcus catus. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Coprococcus comes. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Dorea formicigenerans.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Dorea longicatena.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Enterococcusfaecalis. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isEnterococcus faecium. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Erysipelotrichaceae bacterium 3_1_53. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Escherichia coli. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Escherichia coli S88. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Eubacterium eligens. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Eubacteriumfissicatena. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isEubacterium ramulus. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Eubacterium rectale. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Faecalibacterium prausnitzii. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Flavonifractor plautii. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Fusobacterium mortiferum.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Fusobacteriumnucleatum. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isHoldemania filiformis. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Hydrogenoanaerobacterium saccharovorans. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Klebsiella oxytoca. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Lachnospiraceae bacterium3_1_57FAA_CT1. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isLachnospiraceae bacterium 7_1_58FAA. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Lachnospiraceae bacterium 5_1_57FAA. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Lactobacillus casei. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Lactobacillus rhamnosus. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Lactobacillus ruminis.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Lactococcus casei.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Odoribactersplanchnicus. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isOscillibacter valericigenes. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Parabacteroides gordonii. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Parabacteroides johnsonii. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Parabacteroides merdae. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Pediococcus acidilactici.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Peptostreptococcusasaccharolyticus. In one embodiment, the bacterial entity, e.g., speciesor strain, useful in the compositions and methods of the invention isPropionibacterium granulosum. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Roseburia intestinalis. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Roseburia inulinivorans. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Ruminococcus faecis. In one embodiment,the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Ruminococcus gnavus. In oneembodiment, the bacterial entity, e.g., species or strain, useful in thecompositions and methods of the invention is Ruminococcus sp. ID8. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Ruminococcus torques.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Slackia piriformis.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Staphylococcusepidermidis. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isStaphylococcus saprophyticus. In one embodiment, the bacterial entity,e.g., species or strain, useful in the compositions and methods of theinvention is Streptococcus cristatus. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Streptococcus dysgalactiae subsp. Equisimilis. Inone embodiment, the bacterial entity, e.g., species or strain, useful inthe compositions and methods of the invention is Streptococcus infantis.In one embodiment, the bacterial entity, e.g., species or strain, usefulin the compositions and methods of the invention is Streptococcusoralis. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isStreptococcus sanguinis. In one embodiment, the bacterial entity, e.g.,species or strain, useful in the compositions and methods of theinvention is Streptococcus viridans. In one embodiment, the bacterialentity, e.g., species or strain, useful in the compositions and methodsof the invention is Streptococcus thermophiles. In one embodiment, thebacterial entity, e.g., species or strain, useful in the compositionsand methods of the invention is Veillonella dispar.

In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Acidaminococcus intestine. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Acinetobacter baumannii. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Acinetobacter lwoffii. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Akkermansia muciniphila. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Alistipes putredinis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Alistipes shahii. In one embodiment,the bacterial population useful in the compositions and methods of theinvention comprises Anaerostipes hadrus. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Anaerotruncus colihominis. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Bacteroides caccae. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Bacteroides cellulosilyticus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Bacteroides dorei. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBacteroides eggerthii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBacteroides finegoldii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBacteroides fragilis. In one embodiment, the bacterial population usefulin the compositions and methods of the invention comprises Bacteroidesmassiliensis. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Bacteroides ovatus.In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Bacteroides salanitronis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides salyersiae. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides sp. 1_1_6. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides sp. 3_1_23. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides sp. D20. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides thetaiotaomicrond. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides uniformis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bacteroides vulgatus. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium adolescentis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium bifidum. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium breve. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium faecale. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium kashiwanohense. Inone embodiment, the bacterial population useful in the compositions andmethods of the invention comprises Bifidobacterium longum subsp. Longum.In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Bifidobacteriumpseudocatenulatum. In one embodiment, the bacterial population useful inthe compositions and methods of the invention comprises Bifidobacteriumstercoris. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Blautia(Ruminococcus) coccoides. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBlautia faecis. In one embodiment, the bacterial population useful inthe compositions and methods of the invention comprises Blautiaglucerasea. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Blautia(Ruminococcus) hansenii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBlautia hydrogenotrophica (Ruminococcus hydrogenotrophicus). In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Blautia (Ruminococcus) luti. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Blautia (Ruminococcus) obeum. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Blautia producta (Ruminococcusproductus). In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Blautia(Ruminococcus) schinkii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesBlautia stercoris. In one embodiment, the bacterial population useful inthe compositions and methods of the invention comprises Blautiauncultured bacterium clone BKLE_a03_2 (GenBank: EU469501.1). In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Blautia uncultured bacterium cloneSJTU_B_14_30 (GenBank: EF402926.1). In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Blautia uncultured bacterium clone SJTU_C_14_16 (GenBank:EF404657.1). In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Blautia unculturedbacterium clone S1-5 (GenBank: GQ898099.1). In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Blautia uncultured PACO000178_s(www.ezbiocloud.net/eztaxon/hierarchy?m=browse&k=PAC000178&d=2). In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Blautia wexlerae. In one embodiment,the bacterial population useful in the compositions and methods of theinvention comprises Candidatus arthromitus sp. SFB-mouse-Yit. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Catenibacterium mitsuokai. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridiaceae bacterium (Dielmafastidiosa) JC13. In one embodiment, the bacterial population useful inthe compositions and methods of the invention comprises Clostridialesbacterium 1_7_47FAA. In one embodiment, the bacterial population usefulin the compositions and methods of the invention comprises Clostridiumasparagiforme. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Clostridium bolteae.In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Clostridium clostridioforme. Inone embodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium glycyrrhizinilyticum. Inone embodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium (Hungatella) hathewayi.In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Clostridium histolyticum. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium indolis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium leptum. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium (Tyzzerella) nexile. Inone embodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium perfringens. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Clostridium (Erysipelatoclostridium)ramosum. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Clostridiumscindens. In one embodiment, the bacterial entity, e.g., species orstrain, useful in the compositions and methods of the invention isClostridium septum. In one embodiment, the bacterial population usefulin the compositions and methods of the invention comprises Clostridiumsp. 14774. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Clostridium sp.7_3_54FAA. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Clostridium sp.HGF2. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Clostridiumsymbiosum. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Collinsellaaerofaciens. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Collinsellaintestinalis. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Coprobacillus sp.D7. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Coprococcus catus.In one embodiment, the bacterial population useful in the compositionsand methods of the invention comprises Coprococcus comes. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Dorea formicigenerans. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Dorea longicatena. In one embodiment,the bacterial population useful in the compositions and methods of theinvention comprises Enterococcus faecalis. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Enterococcus faecium. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Erysipelotrichaceae bacterium 3_1_53. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Escherichia coli. In one embodiment,the bacterial population useful in the compositions and methods of theinvention comprises Escherichia coli S88. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Eubacterium eligens. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Eubacterium fissicatena. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Eubacterium ramulus. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Eubacterium rectale. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Faecalibacterium prausnitzii. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Flavonifractor plautii. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Fusobacterium mortiferum. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Fusobacterium nucleatum. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Holdemania filiformis. In one embodiment, thebacterial population useful in the compositions and methods of theinvention comprises Hydrogenoanaerobacterium saccharovorans. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Klebsiella oxytoca. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Lachnospiraceae bacterium3_1_57FAA_CT1. In one embodiment, the bacterial population useful in thecompositions and methods of the invention comprises Lachnospiraceaebacterium 7_1_58FAA. In one embodiment, the bacterial population usefulin the compositions and methods of the invention comprisesLachnospiraceae bacterium 5_1_57FAA. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Lactobacillus casei. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Lactobacillus rhamnosus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Lactobacillus ruminis. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Lactococcus casei. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesOdoribacter splanchnicus. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesOscillibacter valericigenes. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesParabacteroides gordonii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesParabacteroides johnsonii. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesParabacteroides merdae. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesPediococcus acidilactici. In one embodiment, the bacterial populationuseful in the compositions and methods of the invention comprisesPeptostreptococcus asaccharolyticus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Propionibacterium granulosum. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Roseburia intestinalis. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Roseburia inulinivorans. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Ruminococcus faecis. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Ruminococcus gnavus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Ruminococcus sp. ID8. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Ruminococcus torques. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Slackia piriformis. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Staphylococcus epidermidis. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Staphylococcus saprophyticus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Streptococcus cristatus. In one embodiment, the bacterialpopulation useful in the compositions and methods of the inventioncomprises Streptococcus dysgalactiae subsp. Equisimilis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Streptococcus infantis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Streptococcus oralis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Streptococcus sanguinis. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Streptococcus viridans. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Streptococcus thermophiles. In oneembodiment, the bacterial population useful in the compositions andmethods of the invention comprises Veillonella dispar.

In some embodiments, the pharmaceutical composition comprises engineeredbacteria. For example, engineered bacteria include bacteria harboring i)one or more genetic changes, such change being an insertion, deletion,translocation, or substitution, or any combination thereof, of one ormore nucleotides contained on the bacterial chromosome or on anendogenous plasmid, wherein the genetic change may result in thealteration, disruption, removal, or addition of one or moreprotein-coding genes, non-protein-coding genes, gene regulatory regions,or any combination thereof, and wherein such change may be a fusion oftwo or more separate genomic regions or may be synthetically derived;ii) one or more foreign plasmids containing a mutant copy of anendogenous gene, such mutation being an insertion, deletion, orsubstitution, or any combination thereof, of one or more nucleotides;and iii) one or more foreign plasmids containing a mutant or non-mutantexogenous gene or a fusion of two or more endogenous, exogenous, ormixed genes. The engineered bacteria may be produced using techniquesincluding but not limited to site-directed mutagenesis, transposonmutagenesis, knock-outs, knock-ins, polymerase chain reactionmutagenesis, chemical mutagenesis, ultraviolet light mutagenesis,transformation (chemically or by electroporation), phage transduction,or any combination thereof. Suitable bacteria for engineering are knownin the art. For example, as described in PCT Publications Nos. WO93/18163, DELIVERY AND EXPRESSION OF A HYBRID SURFACE PROTEIN ON THESURFACE OF GRAM POSITIVE BACTERIA; WO 03/06593, METHODS FOR TREATINGCANCER BY ADMINISTERING TUMOR-TARGETED BACTERIA AND AN IMMUNOMODULATORYAGENT; and WO 2010/141143, ENGINEERED AVIRULENT BACTERIA STRAINS AND USEIN MEDICAL TREATMENTS.

In some embodiments, the engineered bacteria are natural humancommensals. In other embodiments, the engineered bacteria are attenuatedstrains of pathogens, and may include, but are not limited to,Pseudomonas aeruginosa, Salmonella species, Listeria monocytogenes,Mycoplasma hominis, Escherichia coli, Shigella species, andStreptococcus species, see, e.g. PCT Publications No. WO 03/06593,METHODS FOR TREATING CANCER BY ADMINISTERING TUMOR-TARGETTED BACTERIAAND AN IMMUNOMODULATORY AGENT. Attenuated strains of pathogens will lackall or parts of virulence operons, may lack immune-stimulatory surfacemoieties (e.g., lipopolysaccharide for Gram-negative bacteria), or maycontain one or more nutrient auxotrophies. In specific embodiments, theengineered bacteria are attenuated intracellular pathogens, such asavirulent strains of Listeria monocytogenes.

In some embodiments, the composition of the invention comprises one ormore types of bacteria (e.g., one or more bacterial species or more thanone strain of a particular bacterial species) capable of producingbutyrate in a mammalian subject. Butyrate-producing bacteria may beidentified experimentally, such as by NMR or gas chromatography analysesof microbial products or colorimetric assays (Rose (1955) METHODSENZYMOL. 1: 591-5). Butyrate-producing bacteria may also be identifiedcomputationally, such as by the identification of one or more enzymesinvolved in butyrate synthesis. Non-limiting examples of enzymes foundin butyrate-producing bacteria include butyrate kinase,phosphotransbutyrylase, and butyryl CoA:acetate CoA transferase (Louiset al. (2004) J. BACT. 186(7): 2099-2106). Butyrate-producing strainsinclude, but are not limited to, Faecalibacterium prausnitzii,Eubacterium spp., Butyrivibrio fibrisolvens, Roseburia intestinalis,Clostridium spp., Anaerostipes caccae, and Ruminococcus spp. In someembodiments, a pharmaceutical composition comprises two or more types ofbacteria (e.g., two or more bacterial species or two or more strains ofa particular bacterial species), wherein at least two types of bacteriaare capable of producing butyrate in a mammalian subject. In otherembodiments, the pharmaceutical composition comprises two or more typesof bacteria, wherein two or more types of bacteria cooperate (i.e.,cross-feed) to produce an immunomodulatory SCFA (e.g., butyrate) in amammalian subject. In a preferred embodiment, the pharmaceuticalcomposition comprises at least one type of bacteria (e.g.,Bifidobacterium spp.) capable of metabolizing a prebiotic, including butnot limited to, inulin, inulin-type fructans, or oligofructose, suchthat the resulting metabolic product may be converted by a second typeof bacteria (e.g., a butyrate-producing bacteria such as Roseburia spp.)to an immunomodulatory SCFA such as butyrate (see, e.g., Falony et al.(2006) APPL. ENVIRON. MICROBIOL. 72(12): 7835-7841). In other aspects,the composition comprises at least one acetate-producing bacteria (e.g.,Bacteroides thetaiotaomicron) and at least one acetate-consuming,butyrate-producing bacteria (e.g., Faecalibacterium prausnitzii).

In some embodiments, the pharmaceutical composition comprises one ormore types of bacteria (e.g., one or more bacterial species or more thanone strain of a particular bacterial species) capable of producingpropionate in a mammalian subject, optionally further comprising aprebiotic or substrate appropriate for proprionate biosynthesis.Examples of prebiotics or substrates used for the production ofpropionate include, but are not limited to, L-rhamnose, D-tagalose,resistant starch, inulin, polydextrose, arabinoxylans, arabinoxylanoligosaccharides, mannooligosaccharides, and laminarans (Hosseini et al.(2011) NUTRITION REVIEWS 69(5): 245-258). Propionate-producing bacteriamay be identified experimentally, such as by NMR or gas chromatographyanalyses of microbial products or colorimetric assays (Rose (1955)).Propionate-producing bacteria may also be identified computationally,such as by the identification of one or more enzymes involved inpropionate biosynthesis. Non-limiting examples of enzymes found inpropionate-producing bacteria include enzymes of the succinate pathway,including but not limited to phophoenylpyrvate carboxykinase, pyruvatekinase, pyruvate carboxylase, malate dehydrogenase, fumarate hydratase,succinate dehydrogenase, succinyl CoA synthetase, methylmalonyl Coadecarboxylase, and propionate CoA transferase, as well as enzymes of theacrylate pathway, including but not limited to L-lactate dehydrogenase,propionate CoA transferase, lactoyl CoA dehydratase, acyl CoAdehydrogenase, phosphate acetyltransferase, and propionate kinase.Non-limiting examples of bacteria that utilize the succinate pathway areBacteroides fragilis and other species (including Bacteroides vulgatus),Propionibacterium spp. (including freudenrichii and acidipropionici),Veillonella spp. (including gazogenes), Micrococcus lactilyticus,Selenomonas ruminantium, Escherichia coli, and Prevotella ruminocola.Non-limiting examples of bacteria that utilize the acrylate pathway areClostridium neopropionicum X4, and Megasphaera elsdenii.

In preferred embodiments, the combination of a bacteria and a prebioticis selected based on the fermentation or metabolic preferences of one ormore bacteria capable of producing immunomodulatory SCFAs (e.g.,preference for complex versus simple sugar or preference for afermentation product versus a prebiotic). For example, M. eldseniiprefers lactate fermentation to glucose fermentation, and maximizationof propionate production by M. eldsenii in a mammalian subject maytherefore be achieved by administering along with M. eldsenii a favoredsubstrate (e.g., lactate) or one or more bacteria capable of fermentingglucose into lactate (e.g., Streptococcus bovis) (see, e.g., Hosseini etal. (2011)). Thus, in some embodiments, the composition comprises atleast one type of SCFA-producing bacteria and a sugar fermentationproduct (e.g., lactate). In other embodiments, the composition comprisesat least one type of SCFA-producing bacteria and at least one type ofsugar-fermenting bacteria, wherein the fermentation product of thesecond, sugar-fermenting bacteria is the preferred substrate of theSCFA-producing bacteria.

Immunomodulation can also be achieved by the bacterial production ofglutathione or gamma-glutamylcysteine. Thus, in certain embodiments, thepharmaceutical composition, dosage form, or kit comprises at least onetype of bacteria (e.g., one or more bacterial species or more than onestrain of a particular bacterial species) capable of producingglutathione and/or gamma-glutamylcysteine in a mammalian subject. Insome aspects, the composition comprises one or more bacteria selectedfor the presence of glutamate cysteine ligase (e.g., Lactobacillusfermentum) and/or L-proline biosynthesis enzymes (e.g., E. coli) (Peranet al. (2006) INT. J. COLORECTAL DIS. 21(8): 737-746; Veeravalli et al.(2011) NAT. CHEM. BIO. 7(2): 101-105). In a preferred embodiment, atleast one bacteria in the composition is L. fermentum.

Para-cresol (p-cresol) is a microbial product, via the fermentation oftyrosine or phenylalanine. Sulfated in the liver or colon to p-cresylsulfate, this molecule reduces Th1-mediated responses (Shiba et al.(2014) TOXICOL. APPL. PHARMACOL. 274(2): 191-9). In some embodiments,the composition comprises at least one type of bacteria (e.g., one ormore bacterial species or more than one strain of a particular bacterialspecies) capable of fermenting tyrosine and/or phenylalanine to p-cresolin a mammalian subject. Non-limiting examples of such bacteria includeBacteroides fragilis, Clostridium difficile, and Lactobacillus sp.Strain #11198-(see, e.g., Yokoyama and Carlson (1981) APPL. ENVIRON.MICROBIOL. 41(1): 71-76), and other bacteria with p-hydroxylphenylacetate decarboxylase activity.

In one aspect, provided herein are therapeutic compositions (e.g.,pharmaceutical compositions) containing a purified population ofbacterial cells. As used herein, the terms “purify”, “purified” and“purifying” refer to the state of a population (e.g., a plurality ofknown or unknown amount and/or concentration) of desired bacterialcells, that have undergone one or more processes of purification, e.g.,a selection or an enrichment of the desired bacteria, or alternatively aremoval or reduction of residual habitat products as described herein.In some embodiments, a purified population has no detectable undesiredactivity or, alternatively, the level or amount of the undesiredactivity is at or below an acceptable level or amount. In otherembodiments, a purified population has an amount and/or concentration ofdesired bacterial cells at or above an acceptable amount and/orconcentration. In other embodiments, the purified population ofbacterial cells is enriched as compared to the starting material (e.g.,a fecal material) from which the population is obtained. This enrichmentmay be by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99%, 99.999%, 99.9999%,99.9999%, or greater than 99.999999%, as compared to the startingmaterial.

In certain embodiments, the purified populations of bacterial cells havereduced or undetectable levels of one or more pathogenic activities,such as toxicity, an ability to cause infection of the mammalianrecipient subject, an undesired immunomodulatory activity, an autoimmuneresponse, a metabolic response, or an inflammatory response or aneurological response. Such a reduction in a pathogenic activity may beby 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, 99%, 99.9%, 99.99%, 99.999%, 99.9999%, or greaterthan 99.9999%, as compared to the starting material. In otherembodiments, the purified populations of bacterial cells have reducedsensory components as compared to fecal material, such as reduced odor,taste, appearance, and umami.

In another embodiment, the invention provides purified populations ofbacterial cells that are substantially free of residual habitatproducts. In certain embodiments, this means that the bacterialcomposition no longer contains a substantial amount of the biologicalmatter associated with the microbial community while living on or in thehuman or animal subject, and the purified population of bacterial cells(e.g., bacterial spores or vegetative cells) may be 100% free, 99% free,98% free, 97% free, 96% free, 95% free, 94% free, 93% free, 92% free,91% free, 90% free, 85% free, 80% free, 75% free, 70% free, 60% free, or50% free, of any contamination of the biological matter associated withthe microbial community. Substantially free of residual habitat productsmay also mean that the bacterial composition contains no detectablecells from a human or animal, and that only microbial cells aredetectable, in particular, only desired microbial cells are detectable.In another embodiment, it means that fewer than 1×10⁻²%, 1×10⁻³%,1×10⁻⁴%, 1×10⁻⁵%, 1×10⁻⁶%, 1×10⁻⁷%, 1×10⁻⁸% of the cells in thebacterial composition are human or animal, as compared to microbialcells. In another embodiment, the residual habitat product present inthe purified population is reduced at least a certain level from thefecal material obtained from the mammalian donor subject, e.g., reducedby at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99%, 99.999%, 99.9999%, orgreater than 99.9999%.

In one embodiment, substantially free of residual habitat products orsubstantially free of a detectable level of a pathogenic material meansthat the bacterial composition contains no detectable viral (includingbacterial viruses (i.e., phage)), fungal, or mycoplasmal or toxoplasmalcontaminants, or a eukaryotic parasite such as a helminth.Alternatively, the purified population of bacterial cells (e.g.,bacterial spores and/or vegetative cells) is substantially free of anacellular material, e.g., DNA, viral coat material, or non-viablebacterial material. Alternatively, the purified population of bacterialcells may processed by a method that kills, inactivates, or removes oneor more specific undesirable viruses, such as an enteric virus,including norovirus, poliovirus or hepatitis A virus.

As described herein, purified populations of bacterial cells can bedemonstrated by, for example, genetic analysis (e.g., PCR, DNAsequencing), serology and antigen analysis, microscopic analysis,microbial analysis including germination and culturing, or methods usinginstrumentation such as flow cytometry with reagents that distinguishdesired bacterial entities and/or fungal entities from non-desired,contaminating materials. In yet another embodiment, the spores in apurified population of bacterial cells undergo partial germinationduring processing and formulation such that the final compositioncomprises spores and vegetative bacteria.

In another embodiment, provided herein are methods for production of apharmaceutical composition, e.g., a probiotic composition, comprising apopulation of bacterial cells (e.g., a population of anti-inflammatorybacterial cells), with or without one or more prebiotics, suitable fortherapeutic administration to a mammalian subject in need thereof. Inone embodiment, the composition can be produced by generally followingthe steps of: (a) providing a fecal material obtained from a mammaliandonor subject; and (b) subjecting the fecal material to at least onepurification treatment or step under conditions such that a populationof bacterial cells is produced from the fecal material.

Individual bacterial strains can also be isolated from stool samplesusing culture methods. For example, 5 mls of phosphate-buffered saline(PBS) is added to 1 mg of frozen stool sample and homogenized byvortexing in an anaerobic chamber for isolation of anaerobic bacteria.The suspension is then serially diluted ten-fold (e.g., 10⁻¹ to 10⁻⁹dilutions) and 100 μl aliquots of each dilution are spread evenly overthe surface of agar plates containing different formulations e.g.,anaerobic blood agar plates, Bacteroides bile esculin plates, lakedkanamycin vancomycin plates, egg yolk agar plates and de Man Rogosa andSharpe agar plates. Inverted plates are incubated in an anaerobicchamber for 48 hr+/−4 hours. Colonies with different morphologies arepicked and replated on anaerobic blood agar plates for further testing,PCR analysis and 16 S sequencing. Selected bacterial strains can begrown for therapeutic use singly or in combination.

In one embodiment, a probiotic composition of the invention is not afecal transplant. In some embodiments all or essentially all of thebacterial entities present in a purified population are originallyobtained from a fecal material and subsequently, e.g., for production ofpharmaceutical compositions, are grown in culture as described herein orotherwise known in the art. In some embodiments all or essentially allof the bacterial entities and/or fungal entities present in a purifiedpopulation are obtained from a fecal material and subsequently are grownin culture as described herein or otherwise known in the art. In oneembodiment, the bacterial cells are cultured from a bacterial stock andpurified as described herein. In one embodiment, each of the populationsof bacterial cells are independently cultured and purified, e.g., eachpopulation is cultured separately and subsequently mixed together. Inone embodiment, one or more of the populations of bacterial cells in thecomposition are co-cultured.

Identification of Immunomodulatory Bacteria.

In some embodiments, immunomodulatory bacteria are identified byscreening bacteria to determine whether the bacteria induce secretion ofa pro-inflammatory or a anti-inflammatory cytokines by a host cell(e.g., a host immune cell). In some embodiments, the immunomodulatorybacteria are screened in vitro. For example, human or mammalian cellscapable of cytokine secretion, such as immune cells (e.g., PBMCs,macrophages, T cells, etc.) can be exposed to candidate immunomodulatorybacteria, or supernatants obtained from cultures of candidateimmunomodulatory bacteria, and changes in cytokine expression orsecretion can be measured using standard techniques, such as ELISA,immunoblot, Luminex, antibody array, quantitative PCR, microarray, etc.Bacteria for inclusion in a pharmaceutical composition (e.g., aprobiotic composition) can be selected based on the ability to induce adesired cytokine profile in human or mammalian cells (e.g., immunecells). For example, anti-inflammatory bacteria can be selected forinclusion in a pharmaceutical composition based on the ability to inducesecretion of one or more anti-inflammatory cytokines, and/or the abilityto reduce secretion of one or more pro-inflammatory cytokines.Anti-inflammatory cytokines include, for example, IL-10, IL-13, IL-9,IL-4, IL-5, TGFβ, and combinations thereof. Pro-inflammatory cytokinesinclude, for example, IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1, MIP1α,MIP1β, TNFα, and combinations thereof. In some embodiments,anti-inflammatory bacteria may be selected for inclusion in apharmaceutical compositions based on the ability to modulate thesecretion of one or more anti-inflammatory cytokines and/or that abilityto reduce secretion of one or pro-inflammatory cytokines that have beeninduced by a bacterial cell of a different bacteria type. In someembodiments, the different bacterial cell is of a different bacterialgenus. In some embodiments, the different bacterial cell is of adifferent bacterial species. In some embodiments, the differentbacterial cell is of a different bacterial strain.

In other embodiments, immunomodulatory bacteria are identified byscreening bacteria to determine whether the bacteria impact thedifferentiation and/or expansion of particular subpopulations of immunecells. For example, candidate bacteria can be screened for the abilityto promote differentiation and/or expansion of Treg cells, Th17 cells,Th1 cells and/or Th2 cells from precursor cells, e.g., naive T cells. Byway of example, naïve T cells can be cultured in the presence ofcandidate bacteria or supernatants obtained from cultures of candidatebacteria, and the quantity of Treg cells, Th17 cells, Th1 cells and/orTh2 cells can be determined using standard techniques, such as FACSanalysis. Markers indicative of Treg cells include, for example,CD25⁺CD127^(lo). Markers indicative of Th17 cells include, for example,CXCR3⁻ CCR6⁺. Markers indicative of Th1 cells include, for example,CXCR3⁺CCR6⁻. Markers indicative of Th2 cells include, for example,CXCR3⁻CCR6⁻. Other markers indicative of particular T cellsubpopulations are known in the art, and may be used in the assaysdescribed herein, e.g., to identify populations of immune cells impactedby candidate immunomodulatory bacteria. Bacteria can be selected forinclusion in a pharmaceutical composition based on the ability topromote differentiation and/or expansion of a desired immune cellsubpopulation.

In other embodiments, immunomodulatory bacteria are identified byscreening bacteria to determine whether the bacteria secrete short chainfatty acids (SCFA), such as, for example, butyrate, acetate, propionate,or valerate, or combinations thereof. For example, secretion of shortchain fatty acids into bacterial supernatants can be measured usingstandard techniques. In one embodiment, bacterial supernatants can bescreened to measure the level of one or more short chain fatty acidsusing NMR, mass spectrometry (e.g., GC-MS, tandem mass spectrometry,matrix-assisted laser desorption/ionization, etc.), ELISA, orimmunoblot. Expression of bacterial genes responsible for production ofshort chain fatty acids can also be determined by standard techniques,such as Northern blot, microarray, or quantitative PCR.

In some embodiments, provided herein are pharmaceutical compositionscomprising a population of bacterial cells (e.g., bacterial cells of theorder Clostridiales) containing one type of bacteria. In someembodiments, provided herein are pharmaceutical compositions comprisinga population of bacterial cells (e.g., bacterial cells of the OrderClostridiales) containing more than one type of bacteria. As usedherein, a “type” or more than one “types” of bacteria may bedifferentiated at the genus level, the species level, the sub-specieslevel, the strain level or by any other taxonomic method, as describedherein and otherwise known in the art.

In some embodiment, the pharmaceutical composition may contain one ormore types of bacteria, including bacterial strains of the same speciesor of different species. For instance, a pharmaceutical composition maycomprise bacterial cells of 1, at least 2, at least 3, or at least 4types of bacteria. In another embodiment, a bacterial composition maycomprise at least 5, at least 6, at least 7, at least 8, at least 9, atleast 10, at least 11, at least 12, at least 13, at least 14, at least15, at least 16, at least 17, at least 18, at least 19, or at least 20,at least 30, at least 40, at least 50, or more than 50 types ofbacteria, as defined by species or operational taxonomic unit (OTU)encompassing such species. In a preferred embodiment, a pharmaceuticalcomposition comprises from 2 to no more than 40, from 2 to no more than30, from 2 to no more than 20, from 2 to no more than 15, from 2 to nomore than 10, from 2 to no more than 5, types of bacteria. In anotherpreferred embodiment, a bacterial composition comprises a single type ofbacteria.

In a preferred embodiment, the composition comprises about 20 or fewerisolated populations of bacterial cells. In another embodiment, thecomposition comprises about 15 or fewer isolated populations ofbacterial cells. In another embodiment, the composition comprises about10 or fewer isolated populations of bacterial cells. In anotherembodiment, the composition comprises about 5 or fewer isolatedpopulations of bacterial cells. In another embodiment, the compositioncomprises about 4 or fewer isolated populations of bacterial cells. Inanother embodiment, the composition comprises about 3 or fewer isolatedpopulations of bacterial cells. In another embodiment, the compositioncomprises about 2 isolated populations of bacterial cells. In anotherembodiment, the composition comprises between about 12 and 20 isolatedpopulations of bacterial cells. In another embodiment, the compositioncomprises a single isolated population of bacterial cells. In anotherembodiment, the composition comprises at least two isolated populationsof bacterial cells. In yet another embodiment, the composition comprisesabout 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,or 20 isolated populations of bacterial cells.

In some embodiments, the pharmaceutical composition contains a definedmixture of isolated bacteria. For example, in some embodiments, thepharmaceutical composition contains no more than 100 types of bacteria.In other embodiments, the pharmaceutical composition contains no morethan 75 types of bacteria. In some embodiments, the pharmaceuticalcomposition contains no more than 60 types of bacteria. In otherembodiments, the pharmaceutical composition contains no more than 50types of bacteria. In some embodiments, the pharmaceutical compositioncontains no more than 45 types of bacteria. In other embodiments, thepharmaceutical composition contains no more than 40 types of bacteria.In some embodiments, the pharmaceutical composition contains no morethan 35 types of bacteria. In other embodiments, the pharmaceuticalcomposition contains no more than 30 types of bacteria. In someembodiments, the pharmaceutical composition contains no more than 25types of bacteria. In other embodiments, the pharmaceutical compositioncontains no more than 20 types of bacteria. In some embodiments, thepharmaceutical composition contains no more than 15 types of bacteria.In other embodiments, the pharmaceutical composition contains no morethan 14 types of bacteria. In some embodiments, the pharmaceuticalcomposition contains no more than 13 types of bacteria. In otherembodiments, the pharmaceutical composition contains no more than 12types of bacteria. In some embodiments, the pharmaceutical compositioncontains no more than 11 types of bacteria. In other embodiments, thepharmaceutical composition contains no more than 10 types of bacteria.In some embodiments, the pharmaceutical composition contains no morethan 9 types of bacteria. In other embodiments, the pharmaceuticalcomposition contains no more than 8 types of bacteria. In someembodiments, the pharmaceutical composition contains no more than 7types of bacteria. In other embodiments, the pharmaceutical compositioncontains no more than 6 types of bacteria. In some embodiments, thepharmaceutical composition contains no more than 5 types of bacteria. Inother embodiments, the pharmaceutical composition contains no more than4 types of bacteria. In some embodiments, the pharmaceutical compositioncontains no more than 3 types of bacteria. In other embodiments, thepharmaceutical composition contains no more than 2 types of bacteria. Insome embodiments, the pharmaceutical composition contains no more than 1type of bacteria. In some embodiments, the pharmaceutical compositioncontains defined quantities of each bacterial species. In an exemplaryembodiment, the bacteria incorporated into the pharmaceuticalcomposition are not isolated from fecal matter

Provided herein are pharmaceutical compositions comprising at least one,at least two or at least three types of bacteria that are not identicaland that are capable of decreasing the risk and/or severity of anautoimmune or inflammatory disease, symptom, condition, or disorder, ordysbiosis. In an embodiment, the pharmaceutical composition comprises atleast about 2, 3, 4, 5, 6, 7, 8, 9, or 10 types of isolated bacteria. Inone embodiment, the pharmaceutical composition comprises at least about4 types of isolated bacteria or at least about 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47,48, 49, 50 or more types of isolated bacteria. In some embodiments, theabove invention relates to pharmaceutical compositions furthercomprising one or more prebiotics.

In some embodiments, the pharmaceutical composition of the inventionincludes at least one type of bacteria, wherein said bacteria is abacterial strain (e.g., strain of anti-inflammatory bacterial cells),and the composition includes at least 1×10³ colony forming units (CFU)per dose of said bacterial strain. In other embodiments, thepharmaceutical composition of the invention includes at least one typeof bacteria, wherein said bacteria is a bacterial strain, and thecomposition includes at least about 1×10³, 1×10⁴, 1×10⁵, 1×10⁶, 1×10⁷,1×10⁸, 1×10⁹, 1×10¹⁰, 1×10¹¹, 1×10¹², 1×10¹³, 1×10¹⁴, 1×10¹⁵, or greaterthan 1×10¹⁵ CFU per dose of each bacterial strain present in thecomposition.

In some embodiments, the pharmaceutical compositions of the inventionare formulated for oral or gastric administration, typically to amammalian subject (e.g., a human). In some embodiments, the compositionis formulated for oral administration as a solid, semi-solid, gel, orliquid form, such as in the form of a pill, tablet, capsule, or lozenge.In another embodiment, the pharmaceutical composition is formulated as askin patch. In another embodiment, the pharmaceutical composition isformulated for topical administration. In one embodiment, thepharmaceutical composition is formulated as a food product. In someembodiments, such formulations contain or are coated by an entericcoating to protect the bacterial strain through the stomach and smallintestine, although spores are generally resistant to the stomach andsmall intestines. In other embodiments, the pharmaceutical compositionsmay be formulated with a germinant to enhance engraftment, or efficacy.In yet other embodiments, the pharmaceutical compositions may beco-formulated or co-administered with prebiotic substances, to enhanceengraftment or efficacy.

The composition(s) may include different types of carriers depending onwhether it is to be administered in solid, liquid or aerosol form, andwhether it needs to be sterile for such routes of administration such asinjection. The present invention can be administered intravenously,intradermally, intraarterially, intraperitoneally, intralesionally,intracranially, intraarticularly, intraprostaticaly, intrapleurally,intratracheally, intranasally, intravitreally, intravaginally,intrarectally, topically, intratumorally, intramuscularly,intraperitoneally, subcutaneously, subconjunctival, intravesicularlly,mucosally, intlrapericardially, intraumbilically, intraocularally,orally, topically, locally, as an injection, infusion, continuousinfusion, localized perfusion bathing target cells directly, via acatheter, via a lavage, in lipid compositions (e.g., liposomes), as anaerosol, or by other method or any combination of the fore going aswould be known to one of ordinary skill in the art (see, for example,Remington's Pharmaceutical Sciences, 18th Ed. Mack Printing Company,1990, incorporated herein by reference).

In some embodiments, the composition comprises at least one lipid. Asused herein, a “lipid” includes fats, oils, triglycerides, cholesterol,phospholipids, fatty acids in any form including free fatty acids. Fats,oils and fatty acids can be saturated, unsaturated (cis or trans) orpartially unsaturated (cis or trans). In some embodiments, the lipidcomprises at least one fatty acid selected from lauric acid (12:0),myristic acid (14:0), palmitic acid (16:0), palmitoleic acid (16:1),margaric acid (17:0), heptadecenoic acid (17:1), stearic acid (18:0),oleic acid (18:1), linoleic acid (18:2), linolenic acid (18:3),octadecatetraenoic acid (18:4), arachidic acid (20:0), eicosenoic acid(20:1), eicosadienoic acid (20:2), eicosatetraenoic acid (20:4),eicosapentaenoic acid (20:5) (EPA), docosanoic acid (22:0), docosenoicacid (22:1), docosapentaenoic acid (22:5), docosahexaenoic acid (22:6)(DHA), and tetracosanoic acid (24:0). In other embodiments, thecomposition comprises at least one modified lipid, for example, a lipidthat has been modified by cooking.

In some embodiments, the composition comprises at least one supplementalmineral or mineral source. Examples of minerals include, withoutlimitation: chloride, sodium, calcium, iron, chromium, copper, iodine,zinc, magnesium, manganese, molybdenum, phosphorus, potassium, andselenium. Suitable forms of any of the foregoing minerals includesoluble mineral salts, slightly soluble mineral salts, insoluble mineralsalts, chelated minerals, mineral complexes, non-reactive minerals suchas carbonyl minerals, and reduced minerals, and combinations thereof.

In certain embodiments, the composition comprises at least onesupplemental vitamin. The at least one vitamin can be fat-soluble orwater soluble vitamins. Suitable vitamins include but are not limited tovitamin C, vitamin A, vitamin E, vitamin B12, vitamin K, riboflavin,niacin, vitamin D, vitamin B6, folic acid, pyridoxine, thiamine,pantothenic acid, and biotin. Suitable forms of any of the foregoing aresalts of the vitamin, derivatives of the vitamin, compounds having thesame or similar activity of the vitamin, and metabolites of the vitamin.

In other embodiments, the pharmaceutical composition comprises anexcipient. Non-limiting examples of suitable excipients include abuffering agent, a preservative, a stabilizer, a binder, a compactionagent, a lubricant, a dispersion enhancer, a disintegration agent, aflavoring agent, a sweetener, and a coloring agent.

In another embodiment, the excipient is a buffering agent. Non-limitingexamples of suitable buffering agents include sodium citrate, magnesiumcarbonate, magnesium bicarbonate, calcium carbonate, and calciumbicarbonate.

In some embodiments, the excipient comprises a preservative.Non-limiting examples of suitable preservatives include antioxidants,such as alpha-tocopherol and ascorbate, and antimicrobials, such asparabens, chlorobutanol, and phenol.

In cases where a pharmaceutical composition contains a anaerobicbacterial strain, the pharmaceutical formulation and excipients can beselected to prevent exposure of the bacterial strains to oxygen.

In other embodiments, the composition comprises a binder as anexcipient. Non-limiting examples of suitable binders include starches,pregelatinized starches, gelatin, polyvinylpyrolidone, cellulose,methylcellulose, sodium carboxymethylcellulose, ethylcellulose,polyacrylamides, polyvinyloxoazolidone, polyvinylalcohols, C₁₂-C₁₈ fattyacid alcohol, polyethylene glycol, polyols, saccharides,oligosaccharides, and combinations thereof.

In another embodiment, the composition comprises a lubricant as anexcipient. Non-limiting examples of suitable lubricants includemagnesium stearate, calcium stearate, zinc stearate, hydrogenatedvegetable oils, sterotex, polyoxyethylene monostearate, talc,polyethyleneglycol, sodium benzoate, sodium lauryl sulfate, magnesiumlauryl sulfate, and light mineral oil.

In other embodiments, the composition comprises a dispersion enhancer asan excipient. Non-limiting examples of suitable dispersants includestarch, alginic acid, polyvinylpyrrolidones, guar gum, kaolin,bentonite, purified wood cellulose, sodium starch glycolate,isoamorphous silicate, and microcrystalline cellulose as high HLBemulsifier surfactants.

In some embodiments, the pharmaceutical composition comprises adisintegrant as an excipient. In other embodiments, the disintegrant isa non-effervescent disintegrant. Non-limiting examples of suitablenon-effervescent disintegrants include starches such as corn starch,potato starch, pregelatinized and modified starches thereof, sweeteners,clays, such as bentonite, micro-crystalline cellulose, alginates, sodiumstarch glycolate, gums such as agar, guar, locust bean, karaya, pecitin,and tragacanth. In another embodiment, the disintegrant is aneffervescent disintegrant. Non-limiting examples of suitableeffervescent disintegrants include sodium bicarbonate in combinationwith citric acid, and sodium bicarbonate in combination with tartaricacid.

In another embodiment, the excipient comprises a flavoring agent.Flavoring agents can be chosen from synthetic flavor oils and flavoringaromatics; natural oils; extracts from plants, leaves, flowers, andfruits; and combinations thereof. In some embodiments the flavoringagent is selected from cinnamon oils; oil of wintergreen; peppermintoils; clover oil; hay oil; anise oil; eucalyptus; vanilla; citrus oilsuch as lemon oil, orange oil, grape and grapefruit oil; and fruitessences including apple, peach, pear, strawberry, raspberry, cherry,plum, pineapple, and apricot.

In other embodiments, the excipient comprises a sweetener. Non-limitingexamples of suitable sweeteners include glucose (corn syrup), dextrose,invert sugar, fructose, and mixtures thereof (when not used as acarrier); saccharin and its various salts such as the sodium salt;dipeptide sweeteners such as aspartame; dihydrochalcone compounds,glycyrrhizin; Stevia rebaudiana (Stevioside); chloro derivatives ofsucrose such as sucralose; and sugar alcohols such as sorbitol,mannitol, sylitol, and the like. Also contemplated are hydrogenatedstarch hydrolysates and the synthetic sweetener3,6-dihydro-6-methyl-1,2,3-oxathiazin-4-one-2,2-dioxide, particularlythe potassium salt (acesulfame-K), and sodium and calcium salts thereof.

In yet other embodiments, the composition comprises a coloring agent.Non-limiting examples of suitable color agents include food, drug andcosmetic colors (FD&C), drug and cosmetic colors (D&C), and externaldrug and cosmetic colors (Ext. D&C). The coloring agents can be used asdyes or their corresponding lakes.

The weight fraction of the excipient or combination of excipients in theformulation of the pharmaceutical composition is usually about 99% orless, such as about 95% or less, about 90% or less, about 85% or less,about 80% or less, about 75% or less, about 70% or less, about 65% orless, about 60% or less, about 55% or less, 50% or less, about 45% orless, about 40% or less, about 35% or less, about 30% or less, about 25%or less, about 20% or less, about 15% or less, about 10% or less, about5% or less, about 2% or less, or about 1% or less of the total weight ofthe composition.

The compositions disclosed herein can be formulated into a variety offorms and administered by a number of different means. The compositionscan be administered orally, rectally, or parenterally, in formulationscontaining conventionally acceptable carriers, adjuvants, and vehiclesas desired. The term “parenteral” as used herein includes subcutaneous,intravenous, intramuscular, or intrasternal injection and infusiontechniques. In an exemplary embodiment, the composition is administeredorally.

Solid dosage forms for oral administration include capsules, tablets,caplets, pills, troches, lozenges, powders, and granules. A capsuletypically comprises a core material comprising a bacterial compositionand a shell wall that encapsulates the core material. In someembodiments, the core material comprises at least one of a solid, aliquid, and an emulsion. In other embodiments, the shell wall materialcomprises at least one of a soft gelatin, a hard gelatin, and a polymer.Suitable polymers include, but are not limited to: cellulosic polymerssuch as hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropylmethyl cellulose (HPMC), methyl cellulose, ethyl cellulose, celluloseacetate, cellulose acetate phthalate, cellulose acetate trimellitate,hydroxypropylmethyl cellulose phthalate, hydroxypropylmethyl cellulosesuccinate and carboxymethylcellulose sodium; acrylic acid polymers andcopolymers, such as those formed from acrylic acid, methacrylic acid,methyl acrylate, ammonio methylacrylate, ethyl acrylate, methylmethacrylate and/or ethyl methacrylate (e.g., those copolymers soldunder the trade name “Eudragit”); vinyl polymers and copolymers such aspolyvinyl pyrrolidone, polyvinyl acetate, polyvinylacetate phthalate,vinylacetate crotonic acid copolymer, and ethylene-vinyl acetatecopolymers; and shellac (purified lac). In yet other embodiments, atleast one polymer functions as taste-masking agents.

Tablets, pills, and the like can be compressed, multiply compressed,multiply layered, and/or coated. The coating can be single or multiple.In one embodiment, the coating material comprises at least one of asaccharide, a polysaccharide, and glycoproteins extracted from at leastone of a plant, a fungus, and a microbe. Non-limiting examples includecorn starch, wheat starch, potato starch, tapioca starch, cellulose,hemicellulose, dextrans, maltodextrin, cyclodextrins, inulins, pectin,mannans, gum arabic, locust bean gum, mesquite gum, guar gum, gumkaraya, gum ghatti, tragacanth gum, funori, carrageenans, agar,alginates, chitosans, or gellan gum. In some embodiments the coatingmaterial comprises a protein. In another embodiment, the coatingmaterial comprises at least one of a fat and an oil. In otherembodiments, the at least one of a fat and an oil is high temperaturemelting. In yet another embodiment, the at least one of a fat and an oilis hydrogenated or partially hydrogenated. In one embodiment, the atleast one of a fat and an oil is derived from a plant. In otherembodiments, the at least one of a fat and an oil comprises at least oneof glycerides, free fatty acids, and fatty acid esters. In someembodiments, the coating material comprises at least one edible wax. Theedible wax can be derived from animals, insects, or plants. Non-limitingexamples include beeswax, lanolin, bayberry wax, carnauba wax, and ricebran wax. Tablets and pills can additionally be prepared with entericcoatings.

Alternatively, powders or granules embodying the bacterial compositionsdisclosed herein can be incorporated into a food product. In someembodiments, the food product is a drink for oral administration.Non-limiting examples of a suitable drink include fruit juice, a fruitdrink, an artificially flavored drink, an artificially sweetened drink,a carbonated beverage, a sports drink, a liquid diary product, a shake,an alcoholic beverage, a caffeinated beverage, infant formula and soforth. Other suitable means for oral administration include aqueous andnonaqueous solutions, emulsions, suspensions and solutions and/orsuspensions reconstituted from non-effervescent granules, containing atleast one of suitable solvents, preservatives, emulsifying agents,suspending agents, diluents, sweeteners, coloring agents, and flavoringagents.

In some embodiments, the food product can be a solid foodstuff. Suitableexamples of a solid foodstuff include without limitation a food bar, asnack bar, a cookie, a brownie, a muffin, a cracker, an ice cream bar, afrozen yogurt bar, and the like.

In other embodiments, the pharmaceutical compositions disclosed hereinare incorporated into a therapeutic food. In some embodiments, thetherapeutic food is a ready-to-use food that optionally contains some orall essential macronutrients and micronutrients. In another embodiment,the compositions disclosed herein are incorporated into a supplementaryfood that is designed to be blended into an existing meal. In oneembodiment, the supplemental food contains some or all essentialmacronutrients and micronutrients. In another embodiment, the bacterialcompositions disclosed herein are blended with or added to an existingfood to fortify the food's protein nutrition. Examples include foodstaples (grain, salt, sugar, cooking oil, margarine), beverages (coffee,tea, soda, beer, liquor, sports drinks), snacks, sweets and other foods.

In one embodiment, the formulations are filled into gelatin capsules fororal administration. An example of an appropriate capsule is a 250 mggelatin capsule containing from 10 mg (up to 100 mg) of lyophilizedpowder (10⁸ to 10¹¹ CFUs), 160 mg microcrystalline cellulose, 77.5 mggelatin, and 2.5 mg magnesium stearate. In an alternative embodiment,from 10⁵ to 10¹², 10⁵ to 10⁷, 10⁶ to 10⁷, or 10⁸ to 10¹⁰ CFUs may beused, with attendant adjustments of the excipients if necessary. In analternative embodiment, an enteric-coated capsule or tablet or with abuffering or protective composition can be used.

The pharmaceutical compositions, with or without one or more prebiotics,are generally formulated for oral or gastric administration, typicallyto a mammalian subject. In particular embodiments, the composition isformulated for oral administration as a solid, semi-solid, gel, orliquid form, such as in the form of a pill, tablet, capsule, or lozenge.In some embodiments, such formulations contain or are coated by anenteric coating to protect the bacteria through the stomach and smallintestine, although spores are generally resistant to the stomach andsmall intestines. In other embodiments, the pharmaceutical compositions,with or without one or more prebiotics, may be formulated with agerminant to enhance engraftment, or efficacy. In yet other embodiments,the pharmaceutical compositions may be co-formulated or co-administeredwith prebiotic substances, to enhance engraftment or efficacy. In someembodiments, bacterial compositions may be co-formulated orco-administered with prebiotic substances, to enhance engraftment orefficacy.

In some formulations, the pharmaceutical composition contains at leastabout 0.5%, 1%, 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% orgreater than 90% spores on a mass basis. In some formulations, theadministered dose does not exceed 200, 300, 400, 500, 600, 700, 800, 900milligrams or 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, or 1.9 grams inmass.

The pharmaceutical compositions of the invention may include livemicrobes, dead microbes, microbes that are lyophilized, freeze-dried,and/or substantially dehydrated, or the composition may includebacterial or fungal spores or virions.

Bacterial compositions for use in the pharmaceutical compositions can bedescribed by operational taxonomic units (OTUs). Bacterial compositionsmay be prepared comprising one or at least two types of isolatedbacteria, wherein a first type and a second type are independentlychosen from the species or OTUs listed in Table 1. Additionally, abacterial composition may be prepared comprising at least two types ofisolated bacteria, wherein a first OTU and a second OTU areindependently characterized by, i.e., at least 95%, 96%, 97%, 98%, 99%or including 100% sequence identity to, sequences listed.

Pharmaceutical compositions may be prepared comprising one or at leasttwo types of isolated bacteria, chosen from the species in Table 1,Table 1A, Table 1B, Table 1C, Table 1D, Table 1E, or Table 1F.Generally, the first bacteria and the second bacteria are not the same.The sequences provided in the sequencing listing file for OTUs in Table1 are full 16S sequences. Therefore, in one embodiment, the first and/orsecond OTUs may be characterized by the full 16S sequences of OTUslisted in Table 1. In another embodiment, the first and/or second OTUsmay be characterized by one or more of the variable regions of the 16Ssequence (V1-V9). In some embodiments, at least one of the V1, V2, V3,V4, V5, V6, V7, V8, and V9 regions are used to characterize an OTU. Inone embodiment, the V1, V2, and V3 regions are used to characterize anOTU. In another embodiment, the V3, V4, and V5 regions are used tocharacterize an OTU. In another embodiment, the V4 region is used tocharacterize an OTU.

Using well known techniques, in order to determine the full 16S sequenceor the sequence of any hypervariable region of the 16S sequence, genomicDNA is extracted from a bacterial sample, the 16S rDNA (full region orspecific hypervariable regions) amplified using polymerase chainreaction (PCR), the PCR products cleaned, and nucleotide sequencesdelineated to determine the genetic composition of 16S gene or subdomainof the gene. If full 16S sequencing is performed, the sequencing methodused may be, but is not limited to, Sanger sequencing. If one or morehypervariable regions are used, such as the V4 region, the sequencingmay be, but is not limited to being, performed using the Sanger methodor using a next-generation sequencing method, such as an Illumina(sequencing by synthesis) method using barcoded primers allowing formultiplex reactions.

OTUs can be defined by a combination of nucleotide markers or genes, inparticular highly conserved genes (e.g., “house-keeping” genes), or acombination thereof, full-genome sequence, or partial genome sequencegenerated using amplified genetic products, or whole genome sequence(WGS). Using well defined methods DNA extracted from a bacterial samplewill have specific genomic regions amplified using PCR and sequenced todetermine the nucleotide sequence of the amplified products. In thewhole genome shotgun (WGS) method, extracted DNA will be directlysequenced without amplification. Sequence data can be generated usingany sequencing technology including, but not limited to Sanger,Illumina, 454 Life Sciences, Ion Torrent, ABI, Pacific Biosciences,and/or Oxford Nanopore.

Prebiotics

In one aspect, the pharmaceutical compositions described herein containa prebiotic. In another aspect, the pharmaceutical compositions areco-administered with a prebiotic (e.g., sequentially or concurrently). Aprebiotic is a selectively fermented ingredient that allows specificchanges, both in the composition and/or activity in the gastrointestinalmicrobiota, that confers benefits upon host well-being and health.Prebiotics can include complex carbohydrates, amino acids, peptides, orother nutritional components useful for the survival of the bacterialcomposition. Prebiotics include, but are not limited to, amino acids,biotin, fructooligosaccharide, galactooligosaccharides, inulin,lactulose, mannan oligosaccharides, oligofructose-enriched inulin,oligofructose, oligodextrose, tagatose, trans-galactooligosaccharide,and xylooligosaccharides.

Suitable prebiotics are usually plant-derived complex carbohydrates,oligosaccharides or polysaccharides. Generally, prebiotics areindigestible or poorly digested by humans and serve as a food source forbacteria. Prebiotics which can be used in the pharmaceutical dosageforms, pharmaceutical compositions, and kits provided herein include,without limitation, galactooligosaccharides (GOS),trans-galactooligosaccharides, fructooligosaccharides or oligofructose(FOS), inulin, oligofructose-enriched inulin, lactulose, arabinoxylan,xylooligosaccharides (XOS), mannooligosaccharides, gum guar, gum arabic,tagatose, amylose, amylopectin, xylan, pectin, and the like andcombinations of thereof. Prebiotics can be found in certain foods, e.g.chicory root, Jerusalem artichoke, Dandelion greens, garlic, leek,onion, asparagus, wheat bran, wheat flour, banana, milk, yogurt,sorghum, burdock, broccoli, Brussels sprouts, cabbage, cauliflower,collard greens, kale, radish and rutabaga, and miso. Alternatively,prebiotics can be purified or chemically or enzymatically synthesized.

In some embodiments, the composition comprises at least one prebiotic.In some embodiment, the prebiotic is a carbohydrate. In someembodiments, the composition of the present invention comprises aprebiotic mixture, which comprises at least one carbohydrate. A“carbohydrate” refers to a sugar or polymer of sugars. The terms“saccharide,” “polysaccharide,” “carbohydrate,” and “oligosaccharide”may be used interchangeably. Most carbohydrates are aldehydes or ketoneswith many hydroxyl groups, usually one on each carbon atom of themolecule. Carbohydrates generally have the molecular formula (CH₂O)_(n).A carbohydrate can be a monosaccharide, a disaccharide, trisaccharide,oligosaccharide, or polysaccharide. The most basic carbohydrate is amonosaccharide, such as glucose, sucrose, galactose, mannose, ribose,arabinose, xylose, and fructose. Disaccharides are two joinedmonosaccharides. Exemplary disaccharides include sucrose, maltose,cellobiose, and lactose. Typically, an oligosaccharide includes betweenthree and six monosaccharide units (e.g., raffinose, stachyose), andpolysaccharides include six or more monosaccharide units. Exemplarypolysaccharides include starch, glycogen, and cellulose. Carbohydratescan contain modified saccharide units, such as 2′-deoxyribose wherein ahydroxyl group is removed, 2′-fluororibose wherein a hydroxyl group isreplace with a fluorine, or N-acetylglucosamine, a nitrogen-containingform of glucose (e.g., 2′-fluororibose, deoxyribose, and hexose).Carbohydrates can exist in many different forms, for example,conformers, cyclic forms, acyclic forms, stereoisomers, tautomers,anomers, and isomers. Carbohydrates may be purified from natural (e.g.,plant or microbial) sources (i.e., they are enzymatically synthetized),or they may be chemically synthesized or modified.

Suitable prebiotic carbohydrates can include one or more of acarbohydrate, carbohydrate monomer, carbohydrate oligomer, orcarbohydrate polymer. In certain embodiments, the pharmaceuticalcomposition, dosage form, or kit comprises at least one type of microbeand at least one type of non-digestible saccharide, which includesnon-digestible monosaccharides, non-digestible oligosaccharides, ornon-digestible polysaccharides. In one embodiment, the sugar units of anoligosaccharide or polysaccharide can be linked in a single straightchain or can be a chain with one or more side branches. The length ofthe oligosaccharide or polysaccharide can vary from source to source. Inone embodiment, small amounts of glucose can also be contained in thechain. In another embodiment, the prebiotic composition can be partiallyhydrolyzed or contain individual sugar moieties that are components ofthe primary oligosaccharide (see, e.g., U.S. Pat. No. 8,486,668,PREBIOTIC FORMULATIONS AND METHODS OF USE).

Prebiotic carbohydrates may include, but are not limited tomonosaccharaides (e.g., trioses, tetroses, pentoses, aldopentoses,ketopentoses, hexoses, cyclic hemiacetals, ketohexoses, heptoses) andmultimers thereof, as well as epimers, cyclic isomers, stereoisomers,and anomers thereof. Nonlimiting examples of monosaccharides include (ineither the L- or D-conformation) glyceraldehyde, threose, ribose,altrose, glucose, mannose, talose, galactose, gulose, idose, lyxose,arabinose, xylose, allose, erythrose, erythrulose, tagalose, sorbose,ribulose, psicose, xylulose, fructose, dihydroxyacetone, and cyclic(alpha or beta) forms thereof. Multimers (disaccharides, trisaccharides,oligosaccharides, polysaccharides) thereof include but are not limitedto sucrose, lactose, maltose, lactulose, trehalose, cellobiose,kojibiose, nigerose, isomaltose, sophorose, laminaribiose, gentioboise,turanose, maltulose, palatinose, gentiobiulose, mannobiose, melibiulose,rutinose, rutinulose, xylobiose, primeverose, amylose, amylopectin,starch (including resistant starch), chitin, cellulose, agar, agarose,xylan, glycogen, bacterial polysaccharides such as capsularpolysaccharides, LPS, and peptodglycan, and biofilm exopolysaccharide(e.g., alginate, EPS), N-linked glycans, and O-linked glycans. Prebioticsugars may be modified and carbohydrate derivatives include amino sugars(e.g., sialic acid, N-acetylglucosamine, galactosamine), deoxy sugars(e.g., rhamnose, fucose, deoxyribose), sugar phosphates, glycosylamines,sugar alcohols, and acidic sugars (e.g., glucuronic acid, ascorbicacid).

In some embodiments, the prebiotic carbohydrate component of thepharmaceutical composition, dosage form, or kit consists essentially ofone or more non-digestible saccharides. In one embodiment,non-digestible oligosaccharides the non-digestible oligosaccharides aregalactooligosaccharides (GOS). In another embodiment, the non-digestibleoligosaccharides are fructooligosaccharides (FOS).

In some embodiments, the prebiotic composition of the inventioncomprises one or more of GOS, lactulose, raffinose, stachyose,lactosucrose, FOS (i.e., oligofructose or oligofructan), inulin,isomalto-oligosaccharide, xylo-oligosaccharide, paratinoseoligosaccharide, transgalactosylated oligosaccharides (i.e.,transgalacto-oligosaccharides), transgalactosylate disaccharides,soybean oligosaccharides (i.e., soyoligosaccharides),gentiooligosaccharides, glucooligosaccharides, pecticoligosaccharides,palatinose polycondensates, difructose anhydride III, sorbitol,maltitol, lactitol, polyols, polydextrose, reduced paratinose,cellulose, β-glucose, β-galactose, β-fructose, verbascose, galactinol,and β-glucan, guar gum, pectin, high, sodium alginate, and lambdacarrageenan, or mixtures thereof. The GOS may be a short-chain GOS, along-chain GOS, or any combination thereof. The FOS may be a short-chainFOS, a long-chain FOS, or any combination thereof.

In some embodiments, the prebiotic composition comprises twocarbohydrate species (nonlimiting examples being a GOS and FOS) in amixture of at least 1:1, at least 2:1, at least 5:1, at least 9:1, atleast 10:1, about 20:1, or at least 20:1.

In some embodiments, the prebiotic composition of the inventioncomprises a mixture of one or more non-digestible oligosaccharides,non-digestible polysaccharides, free monosaccharides, non-digestiblesaccharides, starch, or non-starch polysaccharides. In one embodiment, aprebiotic component of a prebiotic composition is a GOS composition. Inone embodiment, a prebiotic composition is a pharmaceutical composition.In one embodiment, a pharmaceutical composition is a GOS composition.

Oligosaccharides are generally considered to have a reducing end and anon-reducing end, whether or not the saccharide at the reducing end isin fact a reducing sugar. Most oligosaccharides described herein aredescribed with the name or abbreviation for the non-reducing saccharide(e.g., Gal or D-Gal), preceded or followed by the configuration of theglycosidic bond (α or β), the ring bond, the ring position of thereducing saccharide involved in the bond, and then the name orabbreviation of the reducing saccharide (e.g., Glc or D-Glc). Thelinkage (e.g., glycosidic linkage, galactosidic linkage, glucosidiclinkage) between two sugar units can be expressed, for example, as 1,4,1→4, or (1-4).

Both FOS and GOS are non-digestible saccharides. R glycosidic linkagesof saccharides, such as those found in, but not limited to, FOS and GOS,make these prebiotics mainly non-digestible and unabsorbable in thestomach and small intestine α-linked GOS (α-GOS) is also not hydrolyzedby human salivary amylase, but can be used by Bifidobacterium bifidumand Clostridium butyricum (Yamashita et al. (2004) J. APPL. GLYCOSCI.51: 115-122). FOS and GOS can pass through the small intestine and intothe large intestine (colon) mostly intact, except where commensalmicrobes and microbes administered as part of a pharmaceuticalcomposition are able to metabolize the oligosaccharides.

GOS (also known as galacto-oligosaccharides, galactooligosaccharides,trans-oligosaccharide (TOS), trans-galacto-oligosaccharide (TGOS), andtrans-galactooligosaccharide) are oligomers or polymers of galactosemolecules ending mainly with a glucose or sometimes ending with agalactose molecule and have varying degree of polymerization (generallythe DP is between 2-20) and type of linkages. In one embodiment, GOScomprises galactose and glucose molecules. In another embodiment, GOScomprises only galactose molecules. In a further embodiment, GOS aregalactose-containing oligosaccharides of the form of[β-D-Gal-(1-6)]_(n)-β-D-Gal-(1-4)-D-Glc wherein n is 2-20. In anotherembodiment, GOS are galactose-containing oligosaccharides of the formGlc α1-4-[β Gal 1-6)]_(n) where n=2-20. In another embodiment, GOS arein the form of α-D-Glc (1-4)-[β-D-Gal-(1-6)-]_(n) where n=2-20. Gal is agalactopyranose unit and Glc (or Glu) is a glucopyranose unit.

In one embodiment, a prebiotic composition comprises a GOS-relatedcompound. A GOS-related compound can have the following properties: a) a“lactose” moiety; e.g., GOS with a gal-glu moiety and any polymerizationvalue or type of linkage; or b) be stimulatory to “lactose fermenting”microbes in the human GI tract; for example, raffinose (gal-fru-glu) isa “related” GOS compound that is stimulatory to both lactobacilli andbifidobacteria.

In one embodiment, a prebiotic composition comprises GOS with a lowdegree of polymerization. In one embodiment a prebiotic compositioncomprising GOS with a low degree of polymerization increases growth ofprobiotic and select commensal bacteria to a greater extent than anequivalent amount of a prebiotic composition comprising GOS with a highdegree of polymerization. In one embodiment, a prebiotic compositioncomprising a high percentage of GOS with a low degree of polymerizationincreases growth of probiotic and beneficial commensal bacteria to agreater extent than an equivalent amount of a prebiotic compositioncomprising a low percentage of GOS with a low degree of polymerization(DP). In one embodiment a prebiotic composition comprises GOS with a DPless than 20, such as less than 10, less than 9, less than 8, less than7, less than 6, less than 5, less than 4, or less than 3. In anotherembodiment a prebiotic composition comprising GOS with a low DPincreases growth of co-formulated or co-administered microbes and/orbeneficial commensal microbes in the GI tract of a subject.

Linkages between the individual sugar units found in GOS and otheroligosaccharides include β-(1-6), β-(1-4), β-(1-3) and β-(1-2) linkages.In one embodiment, the administered oligosaccharides (e.g., GOS) arebranched saccharides. In another embodiment, the administeredoligosacchardies (e.g, GOS) are linear saccharides.

In some embodiments, the GOS comprises a disaccharide Gal α (1-6) Gal,at least one trisaccharide selected from Gal β (1-6)-Gal β (1-4)-Glc andGal β (1-3)-Gal β (1-4)-Glc, the tetrasaccharide Gal β(1-6)-Gal β(1-6)-Gal β (1-4)-Glc and the pentasaccharide Gal β (1-6)-Gal R(1-6)-Gal β (1-6)-Gal β (1-4)-Glc.

In one embodiment, a GOS composition is a mixture of 10 to 45% w/vdisaccharide, 10 to 45% w/v trisaccharide, 10 to 45% w/v tetrasaccharideand 10 to 45% w/v pentasaccharide. In another embodiment, a GOScomposition is a mixture of oligosaccharides comprising 20-28% by weightof R (1-3) linkages, 20-25% by weight of R (1-4) linkages, and 45-55% byweight of R (1-6) linkages. In one embodiment, a GOS composition is amixture of oligosaccharides comprising 26% by weight of 3 (1-3)linkages, 23% by weight of 3 (1-4) linkages, and 51% by weight of R(1-6) linkages.

Alpha-GOS (also called alpha-bond GOS or alpha-linked GOS) areoligosaccharides having an alpha-galactopyranosyl group. Alpha-GOScomprises at least one alpha glycosidic linkage between the saccharideunits. Alpha-GOS are generally represented by α-(Gal)_(n) (n usuallyrepresents an integer of 2 to 10) or α-(Gal). Glc (n usually representsan integer of 1 to 9). Examples include a mixture ofα-galactosylglucose, α-galactobiose, α-galactotriose, α-galactotetraose,and higher oligosaccharides. Additional non-limiting examples includemelibiose, manninootriose, raffinose, stachyose, and the like, which canbe produced from beat, soybean oligosaccharide, and the like.

Commercially available and enzyme synthesized alpha-GOS products arealso useful for the compositions described herein. Synthesis ofalpha-GOS with an enzyme is conducted utilizing the dehydrationcondensation reaction of α-galactosidase with the use of galactose,galactose-containing substance, or glucose as a substrate. Thegalactose-containing substance includes hydrolysates ofgalactose-containing substances, for example, a mixture of galactose andglucose obtained by allowing beta-galactosidase to act on lactose, andthe like. Glucose can be mixed separately with galactose and be used asa substrate with α-galactosidase (see e.g., WO 02/18614). Methods ofpreparing alpha-GOS have been described (see, e.g., EP 1514551 and EP2027863).

In one embodiment, a GOS composition comprises a mixture of saccharidesthat are alpha-GOS and saccharides that are produced bytransgalactosylation using β-galactosidase. In another embodiment, GOScomprises alpha-GOS. In another embodiment, alpha-GOS comprises α-(Gal)₂from 10% to 100% by weight. In one embodiment, GOS comprises onlysaccharides that are produced by transgalactosylation usingβ-galactosidase.

In one embodiment, a GOS composition can comprise GOS with alphalinkages and beta linkages.

In one embodiment, the pharmaceutical composition, dosage form, or kitcomprises, in addition to one or more microbes, an oligosaccharidecomposition that is a mixture of oligosaccharides comprising 1-20% byweight of di-saccharides, 1-20% by weight tri-saccharides, 1-20% byweight tetra-saccharides, and 1-20% by weight penta-saccharides. Inanother embodiment, an oligosaccharide composition is a mixture ofoligosaccharides consisting essentially of 1-20% by weight ofdi-saccharides, 1-20% by weight tri-saccharides, 1-20% by weighttetra-saccharides, and 1-20% by weight penta-saccharides.

In one embodiment, a prebiotic composition is a mixture ofoligosaccharides comprising 1-20% by weight of saccharides with a degreeof polymerization (DP) of 1-3, 1-20% by weight of saccharides with DP of4-6, 1-20% by weight of saccharides with DP of 7-9, and 1-20% by weightof saccharides with DP of 10-12, 1-20% by weight of saccharides with DPof 13-15.

In another embodiment, a prebiotic composition comprises a mixture ofoligosaccharides comprising 50-55% by weight of di-saccharides, 20-30%by weight tri-saccharides, 10-20% by weight tetra-saccharide, and 1-10%by weight penta-saccharides. In one embodiment, a GOS composition is amixture of oligosaccharides comprising 52% by weight of di-saccharides,26% by weight tri-saccharides, 14% by weight tetra-saccharide, and 5% byweight penta-saccharides. In another embodiment, a prebiotic compositioncomprises a mixture of oligosaccharides comprising 45-55% by weighttri-saccharides, 15-25% by weight tetra-saccharides, 1-10% by weightpenta-saccharides.

In certain embodiments, the composition according to the inventioncomprises a mixture of neutral and acid oligosaccharides as disclosed inPCT Application WO 2005/039597 (N.V. Nutricia) and US Patent Application2015/0004130, which are hereby incorporated by reference. In oneembodiment, the acid oligosaccharide has a degree of polymerization (DP)between 1 and 5000. In another embodiment, the DP is between 1 and 1000.In another embodiment, the DP is between 2 and 250. If a mixture of acidoligosaccharides with different degrees of polymerization is used, theaverage DP of the acid oligosaccharide mixture is preferably between 2and 1000. The acid oligosaccharide may be a homogeneous or heterogeneouscarbohydrate. The acid oligosaccharides may be prepared from pectin,pectate, alginate, chondroitine, hyaluronic acids, heparin, heparane,bacterial carbohydrates, sialoglycans, fucoidan, fucooligosaccharides orcarrageenan, and are preferably prepared from pectin or alginate. Theacid oligosaccharides may be prepared by the methods described in PCTApplication WO 01/60378, which is hereby incorporated by reference. Theacid oligosaccharide is preferably prepared from high methoxylatedpectin, which is characterized by a degree of methoxylation above 50%.As used herein, “degree of methoxylation” (also referred to as DE or“degree of esterification”) is intended to mean the extent to which freecarboxylic acid groups contained in the polygalacturonic acid chain havebeen esterified (e.g. by methylation). In some embodiments, the acidoligosaccharides have a degree of methoxylation above about 10%, aboveabout 20%, above about 50%, above about 70%. In some embodiments, theacid oligosaccharides have a degree of methylation above about 10%,above about 20%, above about 50%, above about 70%.

The term neutral oligosaccharides as used in the present inventionrefers to saccharides which have a degree of polymerization of monoseunits exceeding 2, exceeding 3, exceeding 4, or exceeding 10, which arenot or only partially digested in the intestine by the action of acidsor digestive enzymes present in the human upper digestive tract (smallintestine and stomach) but which are fermented by the human intestinalflora and preferably lack acidic groups. The neutral oligosaccharide isstructurally (chemically) different from the acid oligosaccharide. Theterm “neutral oligosaccharides”, as used herein, preferably refers tosaccharides which have a degree of polymerization of the oligosaccharidebelow 60 monose units. The term “monose units” refers to units having aclosed ring structure, e.g., the pyranose or furanose forms. In someembodiments, the neutral oligosaccharide comprises at least 90% or atleast 95% monose units selected from the group consisting of mannose,arabinose, fructose, fucose, rhamnose, galactose, -D-galactopyranose,ribose, glucose, xylose and derivatives thereof, calculated on the totalnumber of monose units contained therein. Suitable neutraloligosaccharides are preferably fermented by the gut flora. Nonlimitingexamples of suitable neutral oligosaccharides are cellobiose(4-O-β-D-glucopyranosyl-D-glucose), cellodextrins((4-O-β-D-glucopyranosyl)n-D-glucose), B-cyclo-dextrins (cyclicmolecules of α-1-4-linked D-glucose; α-cyclodextrin-hexamer,β-cyclodextrin-heptamer and 7-cyclodextrin-octamer), indigestibledextrin, gentiooligosaccharides (mixture of 3-1-6 linked glucoseresidues, some 1-4 linkages), glucooligosaccharides (mixture ofα-D-glucose), isomaltooligosaccharides (linear α-1-6 linked glucoseresidues with some 1-4 linkages), isomaltose(6-O-α-D-glucopyranosyl-D-glucose); isomaltriose(6-O-α-D-glucopyranosyl-(1-6)-α-D-glucopyranosyl-D-glucose), panose(6-O-α-D-glucopyranosyl-(1-6)-α-D-glucopyranosyl-(1-4)-D-glucose),leucrose (5-O-α-D-glucopyranosyl-D-fructopyranoside), palatinose orisomaltulose (6-O-α-D-glucopyranosyl-D-fructose), theanderose(O-α-D-glucopyranosyl-(1-6)-O-α-D-glucopyranosyl-(1-2)-B-D-fructofuranoside), D-agatose, D-lyxo-hexylose, lactosucrose(O-β-D-galactopyranosyl-(1-4)-O-α-D-glucopyranosyl-(1-2)-β-D-fructofuranoside),α-galactooligosaccharides including raffinose, stachyose and other soyoligosaccharides(O-α-D-galactopyranosyl-(1-6)-α-D-glucopyranosyl-β-D-fructofuranoside),β-galactooligosaccharides or transgalacto-oligosaccharides(β-D-galactopyranosyl-(1-6)-[β-D-glucopyranosyl]n-(1-4) α-D glucose),lactulose (4-O-β-D-galactopyranosyl-D-fructose), 4′-galatosyllactose(O-D-galactopyranosyl-(1-4)-O-β-D-glucopyranosyl-(1-4)-D-glucopyranose),synthetic galactooligosaccharide (neogalactobiose, isogalactobiose,galsucrose, isolactose I, II and III), fructans-Levan-type(β-D-(2→6)-fructofuranosyl)_(n) α-D-glucopyranoside),fructans-Inulin-type (β-D-((2→1)-fructofuranosyl)_(n)α-D-glucopyranoside), 1 f-β-fructofuranosylnystose(β-D-((2→1)-fructofuranosyl)n B-D-fructofuranoside),xylooligo-saccharides (B-D-((1→4)-xylose)n, lafinose, lactosucrose andarabinooligosaccharides.

In some embodiments, the neutral oligosaccharide is selected from thegroup consisting of fructans, fructooligosaccharides, indigestibledextrins galactooligo-saccharides (includingtransgalactooligosaccharides), xylooligosaccharides,arabinooligo-saccharides, glucooligosaccharides, mannooligosaccharides,fucooligosaccharides and mixtures thereof.

Suitable oligosaccharides and their production methods are furtherdescribed in Laere K. J. M. (Laere, K. J. M., Degradation ofstructurally different non-digestible oligosaccharides by intestinalbacteria: glycosylhydrolases of Bi. adolescentis. PhD-thesis (2000),Wageningen Agricultural University, Wageningen, The Netherlands), theentire content of which is hereby incorporated by reference.Transgalactooligosaccharides (TOS) are for example sold under thetrademark Vivinal™ (Borculo Domo Ingredients, Netherlands). Indigestibledextrin, which may be produced by pyrolysis of corn starch, comprisesα(1→4) and α(1→6) glucosidic bonds, as are present in the native starch,and contains 1→2 and 1→3 linkages and levoglucosan. Due to thesestructural characteristics, indigestible dextrin containswell-developed, branched particles that are partially hydrolysed byhuman digestive enzymes. Numerous other commercial sources ofindigestible oligosaccharides are readily available and known to skilledpersons in the art. For example, transgalactooligosaccharide isavailable from Yakult Honsha Co., Tokyo, Japan. Soybean oligosaccharideis available from Calpis Corporation distributed by Ajinomoto U.S.A.Inc., Teaneck, N.J.

In a further preferred embodiment, the pharmaceutical compositioncontains a prebiotic mixture of an acid oligosaccharide with a DPbetween 1 and 5000, prepared from pectin, alginate, and mixturesthereof; and a neutral oligosaccharide, selected from the group offructans, fructooligosaccharides, indigestible dextrins,galactooligosaccharides including transgalacto-oligosaccharides,xylooligosaccharides, arabinooligosaccharides, glucooligosaccharides,manno-oligosaccharides, fucooligosaccharides, and mixtures thereof.

In certain embodiments, the prebiotic mixture comprises xylose. In otherembodiments, the prebiotic mixture comprises a xylose polymer (i.e.xylan). In some embodiments, the prebiotic comprises xylose derivatives,such as xylitol, a sugar alcohol generated by reduction of xylose bycatalytic hydrogenation of xylose, and also xylose oligomers (e.g.,xylooligosaccharide). While xylose can be digested by humans, viaxylosyltransferase activity, most xylose ingested by humans is excretedin urine. In contrast, some microorganisms are efficient at xylosemetabolism or may be selected for enhanced xylose metabolism. Microbialxylose metabolism may occur by at least four pathways, including theisomerase pathway, the Weimburg pathway, the Dahms pathway, and, foreukaryotic microorganisms, the oxido-reductase pathway.

The xylose isomerase pathway involves the direct conversion of D-xyloseinto D-xylulose by xylose isomerase, after which D-xylulose isphosphorylated by xylulose kinase to yield D-xylolose-5-phosphate, anintermediate of the pentose phosphate pathway.

In the Weimberg pathway, D-xylose is oxidized to D-xylono-lactone by aD-xylose dehydrogenase. Then D-xylose dehydrogenase is hydrolyzed by alactonase to yield D-xylonic acid, and xylonate dehydratase activitythen yields 2-keto-3-deoxy-xylonate. The final steps of the Weimbergpathway are a dehydratase reaction to form 2-keto glutarate semialdehydeand an oxidizing reaction to form 2-ketoglutarate, an intermediate ofthe Krebs cycle.

The Dahms pathway follows the same mechanism as the Weimberg pathway butdiverges once it has yielded 2-keto-3-deoxy-xylonate. In the Dahmspathway, an aldolase splits 2-keto-3-deoxy-xylonate into pyruvate andglycolaldehyde.

The xylose oxido-reductase pathway, also known as the xylosereductase-xylitol dehydrogenase pathway, begins by the reduction ofD-xylose to xylitol by xylose reductase followed by the oxidation ofxylitol to D-xylulose by xylitol dehydrogenase. As in the isomerasepathway, the next step in the oxido-reductase pathway is thephosphorylation of D-xylulose by xylulose kinase to yieldD-xylolose-5-phosphate.

Xylose is present in foods like fruits and vegetables and other plantssuch as trees for wood and pulp production. Thus, xylose can be obtainedin the extracts of such plants. Xylose can be obtained from variousplant sources using known processes including acid hydrolysis followedby various types of chromatography. Examples of such methods to producexylose include those described in Maurelli, L. et al. (2013), Appl.Biochem. Biotechnol. 170:1104-1118; Hooi H. T et al. (2013), Appl.Biochem. Biotechnol. 170:1602-1613; Zhang H-J. et al. (2014), BioprocessBiosyst. Eng. 37:2425-2436.

Preferably, the metabolism of xylose and/or the shift in microbiota dueto the metabolism of the xylose provided in a pharmaceutical compositionof the invention confers a benefit to a host, e.g. immunologicaltolerance. For example, in aspects in which the patient is at risk orsuffering from GVHD, the immunological tolerance may reducegraft-versus-host activity while maintaining graft-versus-leukemiaactivity. In another example, in aspects in which the patient suffersfrom Celiac disease, the immunological tolerance prevents aninappropriate immune response to gluten. The xylose may be, e.g. i)cytotoxic for an autoimmune disease- and/or inflammatorydisease-associated associated pathogen or pathobiont, ii) cytostatic foran autoimmune disease- and/or inflammatory disease-associated pathogenor pathobiont, iii) capable of decreasing the growth of autoimmunedisease- and/or inflammatory disease-associated pathogen or pathobiont,iv) capable of inhibiting the growth of an autoimmune disease- and/orinflammatory disease-associated pathogen or pathobiont, v) capable ofdecreasing the colonization of an autoimmune disease- and/orinflammatory disease-associated pathogen or pathobiont, vi) capable ofinhibiting the colonization of an autoimmune disease- and/orinflammatory disease-associated pathogen or pathobiont, vii) capable ofeliciting an immunomodulatory response in the host that reduces the riskof an autoimmune and/or inflammatory disorder, viii), capable ofeliciting an immunomodulatory response in the host that reduces theseverity of an autoimmune and/or inflammatory disorder, ix) capable ofpromoting barrier integrity directly or indirectly through its impact onmicrobiota, or x) any combination of i)-ix).

In some embodiments, the pharmaceutical composition or dosage formcomprises a bacterial population and xylose in an amount effective topromote the growth of select bacteria of the family Clostridiacea,including members of the genus Clostridium, Ruminococcus, or Blautia orrelatives thereof in a host. In some embodiments, the pharmaceuticalcomposition or dosage form is further effective to promote theproliferation of select bacteria of the family Clostridiacea, includingmembers of the genus Clostridium, Ruminococcus, or Blautia or relativesthereof in a host. In certain embodiments, the pharmaceuticalcomposition or dosage form comprises a bacterial population and xylosein an amount effective to promote the colonization and/or engraftment ofselect bacteria of the family Clostridiacea, including members of thegenus Clostridium, Ruminococcus, or Blautia or relatives thereof in ahost. In preferred embodiments, the pharmaceutical composition or dosageform is further capable of altering a dysbiotic state such that thegrowth, proliferation, colonization, and/or engraftment of a host by apathogen, pathobiont, disease-associated microbe, or a combinationthereof such that the population of at least one pathogen, pathobiont,or disease-associated microbe is decreased 2-fold, 5-fold, 10-fold,50-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 10000-fold, or over10000-fold. In one embodiment, the pharmaceutical composition or dosageform is capable of locally or systemically eliminating at least onepathogen, pathobiont, or disease-associated microbe from a host.

In some embodiments, the prebiotic comprises a carbohydrate monomer orpolymer that has been modified i.e., substituted with other substituents(e.g., acetyl group, glucuronic acid residue, arabinose residue, or thelike) (see US Patent Application 20090148573, hereby incorporated byreference). The term “modified”, as used herein, refers to a moleculemodified from a reference molecule, and includes not only artificiallyproduced molecules but also naturally occurring molecules. In preferredembodiments, the modification occurs at one or more hydroxyl groups ofthe reference carbohydrate. In some embodiments, the modification occursat carbon-2 (C2), the modification occurs at carbon-6 (C6), or acombination thereof.

In some embodiments, a carbohydrate (a monomer or, preferably, apolymer) is modified with one or more hydrophilic groups. Nonlimitingexamples of the hydrophilic groups include an acetyl group, a4-O-methyl-α-D-glucuronic acid residue, an L-arabinofuranose residue, anL-arabinose residue, and an α-D-glucuronic acid residue. In someembodiments, the modification is the replacement of one or more hydroxylgroups with —H, —CH₂OH, —CH₃, or —NH₂.

In some embodiments, the composition comprises at least one carbohydratethat elicits an immunomodulatory response. Exemplary immunomodularycarbohydrates include (but are not limited to) fructo-oligosaccharides,glycosaminoglycans (e.g., heparin sulfate, chondroitin sulfate A,hyaluronan), O-glycans, and carrageenan oligosaccharides, andgalacto-oligosaccharides. Immunomodulatory carbohydrates may be purifiedfrom plants or microbes or may be synthetically derived.Immunomodulatory carbohydrates may be effective to, for example, preventdisease, suppress symptoms, treat disease, or any combination thereof.

In some embodiments, immunomodulatory carbohydrates are C-type lectinreceptor ligands. In preferred embodiments, the C-type lectin receptorligands are produced by one or more fungal species. In otherembodiments, the immunomodulatory carbohydrates are bacterialexopolysaccharides, such as (but not limited to) the exopolysaccharides(EPS) produced by Bacillus subtilis, Bifidobacterium breve, orBacteroides fragilis. In some aspects, immunomodulatory carbohydratesare zwitterionic polysaccharides. In some aspects, immunomodulatorycarbohydrates modulate toll-like receptor 2 (TLR2) and/or toll-likereceptor 4 (TLR4) responses in a host. For example, autoimmune orinflammatory diseases characterized by intestinal inflammation may beprevented by a TLR4 agonist such as but not limited to B. subtilis EPS(see, e.g., Jones et al. (2014) J. IMMUNOL. 192: 4813-4820).Immunomodulatory carbohydrates may also activate CD4+ T cells and/orlead to an upregulation of the anti-inflammatory cytokine interleukin-10(Mazmanian and Kasper (2006) NAT. REV. IMMUNOL. 6: 849-858).Immunomodulatory carbohydrates may be selected for administration to apatient based on the presence, abundance, distribution, modificationand/or linkages of sugar residues. For example, immunomodulatorycarbohydrates used in the prevention of intestinal disorders orautoimmune conditions that manifest in the gut (non-limiting examplesbeing IBD and GVHD) may be selected based on i) a high abundance ofmannose residues; ii) the presence of terminal mannopyransosyl (t-Man)residues and/or 2,6 linked mannopyranosyl residues (2,6-Man), iii) aratio of mannose to glucose residues in the approximate range of 8:2 to9:1, iv) the presence of galactose residues, v) areas of positivecharge, or vi) a combination thereof.

Carbohydrates may be selected according to the fermentation or metabolicpreferences of a microbe (e.g., an anti-inflammatory bacterial cell)selected for administration to a mammalian subject. Selection criteriainclude but are not limited to sugar complexity (e.g., monosaccharides,including but not limited to glucose, versus oligosaccharides orstarches) as well as by desired end-product. Non-liming examples includethe fermentation products ethanol and carbon dioxide (CO₂) (e.g., viaethanol fermentation by Saccharomyces sp. Zymomonas sp.), lactate (e.g.,via homolactic acid fermentation by Lactococcus sp., Streptococcus sp.,Enterococcus sp., Pediococcus sp. and some species Lactobacillus),lactate, ethanol, and CO₂ (e.g., via heterolactic acid fermentation(which includes the phosphoketolase pathway) by some species ofLactobacillus as well as Leuconostoc sp., Oenococcus sp., and Weissellasp.), butanol, acetone, CO₂ and H₂ (via acetone-butanol fermentation bysome Clostridium sp.), and short chain fatty acids (with or without theproduction of other products) (see, e.g., Muller (2011) BacterialFermentation. Encyclopedia of Life Sciences). Examples of fermentationleading to short chain fatty acid production include homoacetic acidfermentation (e.g., by Acetobacterium sp., and resulting in acetate),propionic acid fermentation (e.g., by Propionibacterium sp., andresulting in propionate, acetate and CO₂) mixed acid fermentation (e.g.,by Escherichia sp., and resulting in ethanol, lactate, acetate,succinate, formate, CO₂, and H₂), butyrate fermentation (e.g., by someClostridium sp., resulting in butyrate, CO₂, and H₂), and 2,3-butanediolfermentation (e.g., by Enterobacter sp., resulting in ethanol,butanediol, lactate, formate, CO₂, and H₂). In some embodiments,selection of carbohydrates for co-formulation of co-administration witha type of microbe or types of microbe may be achieved by computationalanalysis of microbial enzymatic pathways, including but not limited tothe presence of metabolic/fermentation pathway enzymes.

Other prebiotics include molecules capable of selective orsemi-selective utilization by microbes (e.g., bacterial cells) of thecompositions contained herein. The ability of a microbe to utilize ametabolite of interest is determined by the genomic capacity of thatmicrobe. Public databases have characterized many microbes and automatethe annotation of the genome to allow a computational analysis of themetabolites a microbe is potentially able to utilize. Databases such asthe Cluster of Orthologous Groups (COGs) database characterize genomesfrom a variety of species in this manner and are capable ofcharacterizing newly sequenced genomes as well (e.g. see in this fashion(Tatusov et al. (2000) NUCL. ACID RES. 28(1): 33-36). Furthermore,pathway analysis classifies COGs into different categories withassociated one letter codes including J, translation; L replication,recombination, and repair, K transcription; 0 molecular chaperones andrelated functions, M, cell wall structure and biogenesis and outermembrane, N secretion motility and chemotaxis; T signal Transduction; Pinorganic ion transport and metabolism; C energy production andconversion; G, carbohydrate metabolism and transport; E amino acidmetabolism and transport; F, nueclotide metabolism and transport; D cellDivision and chromosome partitioning; R general functional prediction.In preferred embodiments, COGs of the categories, N, M, P, C, G, E, andF are selected as preferred COGs to both provide enhanced growth onspecific substrates and modified behaviors relevant for anti-tumorproperties.

COGs are selected to be specific or semi enriched in the host or othermicrobes within a host by searching for specific functions present inthe microbe of interest but absent from a large set of other competitionorganisms. Tissue specific analysis of the host for enzymes expressedwithin a tissue is performed to identify tissue specific enzymaticactivities in the host. Specific functions are absent from at least 90%,at least 80%, at least 70%, at least 60%, at least 50%, at least 40%, atleast 30% at least 20% or at least 10% of the other organisms selectedfrom the group of the host, the host tissue, the disease-associatedmicrobiota, the host gut microbiota, the host niche specific to theengraftment of the microbial composition (e.g., GI tract, skin).

Once these COGs are identified, databases like KEGG are used to link theenzymatic functions to identify the metabolites that are substrates forthese selective COGs. Furthermore, the selective analysis to generateselective metabolites is repeated on the set of substrate of COGs tovalidate that the pathways and metabolites are selective to the desiredmicrobial composition.

Methods of the Invention

In one aspect, the invention provides methods for modulating an immuneresponse in a subject in need thereof, the method comprisingadministering a pharmaceutical composition of the invention to therebymodulate the immune response in the subject. In some embodiments, theimmune response is against a microorganism. In some embodiments, theimmune response is against self (e.g., an auto-immune response). In someembodiments, the immune response is a pro-inflammatory immune response.

In another aspect, the invention provides methods for reducinginflammation in a subject in need thereof, the method comprisingadministering a pharmaceutical composition of the invention to therebyreduce inflammation in the subject. In some embodiments, the immuneresponse is against a microorganism. In some embodiments, the subjecthas an autoimmune or inflammatory disorder. In some embodiments, theautoimmune or inflammatory disorder is selected from the groupconsisting of graft-versus-host disease (GVHD), an inflammatory boweldisease (IBD), ulterative colitis, Crohn's disease, multiple sclerosis(MS), systemic lupus erythematosus (SLE), type I diabetes, rheumatoidarthritis, Sjögren's syndrome, and Celiac disease. In an exemplaryembodiments, the autoimmune or inflammatory disorder is GVHD. In anotherexemplary embodiment, the autoimmune or inflammatory disorder is IBD. Inyet another exemplary embodiment, the autoimmune or inflammatorydisorder is ulcerative colitis. In an exemplary embodiment, theautoimmune or inflammatory disorder is Crohn's disease. In anotherexemplary embodiment, the autoimmune or inflammatory disorder ismultiple sclerosis (MS). In yet another embodiment, the autoimmune orinflammatory disorder is systemic lupus erythematosus. In an exemplaryembodiment, the autoimmune or inflammatory disorder is type I diabetes.In another exemplary embodiment, the autoimmune or inflammatory disorderis rheumatoid arthritis. In yet another exemplary embodiment, theautoimmune or inflammatory disorder is rheumatoid arthritis. In anexemplary embodiment, the autoimmune or inflammatory disorder isSjögren's syndrome. In another exemplary embodiment, the autoimmune orinflammatory disorder is Celiac disease.

Autoimmune and inflammatory diseases that may be treated with thepharmaceutical compositions of the present invention, include, but arenot limited to: Acute Disseminated Encephalomyelitis, Acute necrotizinghemorrhagic leukoencephalitis, Addison's disease, adhesive capsulitis,Agammaglobulinemia, Alopecia areata, Amyloidosis, Ankylosingspondylitis, Anti-GBM nephritis, Anti-TBM nephritis, Antiphospholipidsyndrome, arthofibrosis, atrial fibrosis, autoimmune angioedema,autoimmune aplastic anemia, autoimmune dusautonomia, autoimmunehepatitis, autoimmune hyperlipidemia, autoimmune immunodeficiency,autoimmune inner ear disease, autoimmune myocarditis, autoimmuneoophoritis, autoimmune pancreatitis, autoimmune retinopathy, autoimmunethrombocytopenic purpura, autoimmune thyroid disease, autoimmuneurticaria, axonal and neuronal neuropathies, Balo disease, Behçet'sdisease, benign mucosal pemphigold, Bullous pemphigold, cardiomyopathy,Castleman disease, Celiac Disease, Chagas disease, chronic fatiguesyndrome, chronic inflammatory demyelinating polyneuropathy, chronicLyme disease, chronic recurrent multifocal osteomyelitis, Churg-Strausssyndrome, cicatricial pemphigold, cirrhosis, Cogans syndrome, coldagglutinin disease, congenital heart block, Coxsackle myocarditis, CRESTdisease, Crohn's disease, Cystic Fibrosis, essential mixedcryoglobulinemia, deficiency of the interleukin-1 receptor antagonist,demyelinating neuropathies, dermatitis herpetiformis, dermatomyosis,Devic's disease, discoid lupus, Dressler's syndrome, Dupuytren'scontracture, endometriosis, endomyocardial fibrosis, eosinophilicesophagitis, eosinophilic facsciitis, erythema nodosum, experimentalallergic encephalomyelitis, Evans syndrome, Familial MediterraneanFever, fibromyalgia, fibrosing alveolitis, giant cell arteritis, giantcell myocarditis, glomerulonephritis, Goodpasture's syndrome,Graft-versus-host disease (GVHD), granulomatosus with polyanglitis,Graves' disease, Guillain-Bare syndrome, Hashimoto's encephalitis,Hashimoto's thyroiditis, hemolytic anemia, Henoch-Schonlein purpura,hepatitis, herpes gestationis, hypogammaglobulinemia, idiopathicthrombocytopenic purpura, IgA nephropathy, IgG4-related sclerosingdisease, immunoregulatory lipoproteins, inclusion body myositis,inflammatory bowel disorders, interstitial cystitis, juvenile arthritis,juvenile myositis, Kawasaki syndrome, keloid, Lambert-Eaton syndrome,leukocytoclastic vasculitis, lichen planus, lichen sclerosus, ligneousconjunctivitis, linear IgA disease, mediastinal fibrosis, Meniere'sdisease, microscopic polyanglitis, mixed connective tissue disease,Mooren's ulcer, Mucha-Hamermann disease, Multiple Sclerosis (MS),Myasthenia gravis, myelofibrosis, Myositis, narcolepsy, Neonatal OnsetMultisystem Inflammatory Disease, nephrogenic systemic fibrosis,neutropenia, nonalcoholic fatty liver disease, nonalcoholicsteatohepatitis (NASH), ocular-cicatricial pemphigold, optic neuritis,palindromic rheumatism, Pediatric Autoimmune Neuropsychiatric DisordersAssociated with Streptococcus (PANDAS), paraneoplastic cerebellardegeneration, paroxysmal nocturnal nemoglobinuria, Parry Rombergsyndrome, Parsonnage-Turner syndrome, Pars planitis, Pemphigus,Peripheral neuropathy, perivenous encephalomyelitis, pernicious anemia,Peyronie's disease, POEMS syndrome, polyarteritis nodosa, progressivemassive fibrosis, Tumor Necrosis Factor Receptor-associated PeriodicSyndrome, Type I autoimmune polyglandular syndrome, Type II autoimmunepolyglandular syndrome, Type III autoimmune polyglandular syndrome,polymyalgia rhematica, polymyositis, postmyocardial infarction syndrome,postpericardiotomy syndrome, progesterone dermatitis, primary biliarycirrhosis, primary sclerosing cholangitis, psoriasis, psoriaticarthritis, idiopathic pulmonary fibrosis, pyoderma gangrenosum, pure redcell aplasia, Raynauds phenomenon, reactic arthritis, reflex sympatheticdystrophy, Reiter's syndrome, relapsing polychondritis, restless legssyndrome, retroperitoneal fibrosis, rheumatic fever, rheumatoidarthritis, sarcoidosis, Schmidt syndrome, scleritis, scleroderma,Sjögren's syndrome, sperm and testicular autoimmunity, stiff personsyndrome, subacute bacterial endocarditis, Susac's syndrome, sympatheticophthalmia, systemic lupus erythematosus (SLE), Takayasu's arthritis,temporal arteritis, thrombocytopenic purpura, Tolosa-Hunt syndrome,transverse myelitis, Type 1 diabetes, ulcerative colitis,undifferentiated connective tissue disease, uveitis, vasculitis,vesiculobullous dermatosis, and Vitiligo.

The microbes described herein may additively or synergistically reducethe number of types of autoimmune disease- or inflammatorydisease-associated pathogens or pathobionts either distally—e.g.,orally-administered microbes reduce the total microbial burden in anorgan not in the gastrointestinal tract, or intravaginally-administeredmicrobes reduce the total microbial burden in an organ that is not thevagina—or locally, e.g., the intestines or vagina, respectively. Distalsites include but are not limited to the liver, spleen, fallopian tubesand uterus.

Similarly, the microbes described herein may additively orsynergistically elicit an immunomodulatory response either distally,e.g., in which enteral administration of microbes results in alteringthe immune response at the skin or liver, or locally, e.g., the enteraladministration of microbes results in altering the immune response inthe intestines.

In some situations, the recipient subject is immunocompromised orimmunosuppressed, or is at risk of developing an immune or inflammatorydisorder.

In embodiments, the microbial composition is administered enterically,with or without prebiotics. This preferentially includes oraladministration, or by an oral or nasal tube (including nasogastric,nasojejunal, oral gastric, or oral jejunal). In other embodiments,administration includes rectal administration (including enema,suppository, or colonoscopy). The pharmaceutical composition may beadministered to at least one region of the gastrointestinal tract,including the mouth, esophagus, stomach, small intestine, largeintestine, and rectum. In some embodiments, the pharmaceuticalcomposition is administered to all regions of the gastrointestinaltract. The pharmaceutical compositions may be administered orally in theform of medicaments such as powders, capsules, tablets, gels or liquids.The pharmaceutical compositions may also be administered in gel orliquid form by the oral route or through a nasogastric tube, or by therectal route in a gel or liquid form, by enema or instillation through acolonoscope or by a suppository. In some embodiments, the pharmaceuticalcomposition of the invention is administered enterically with one oremore prebiotics.

If the composition is administered colonoscopically and, optionally, ifthe microbial composition, with or without one or more prebiotics, isadministered by other rectal routes (such as an enema or suppository) oreven if the subject has an oral administration, the subject may have acolonic-cleansing preparation. The colon-cleansing preparation canfacilitate proper use of the colonoscope or other administrationdevices, but even when it does not serve a mechanical purpose it canalso maximize the proportion of bacteria from the pharmaceuticalcomposition relative to the other organisms previously residing in thegastrointestinal tract of the subject. Any ordinarily acceptablecolonic-cleansing preparation may be used such as those typicallyprovided when a subject undergoes a colonoscopy.

Pretreatment protocols. Prior to administration of the pharmaceuticalcomposition, with or without one or more prebiotics, the subject canoptionally have a pretreatment protocol to prepare the gastrointestinaltract or vagina to receive the pharmaceutical composition. In theseinstances, the pretreatment protocol can enhance the ability of thepharmaceutical composition to affect the patient's microbiome.

As one way of preparing the patient for administration of the microbialecosystem, at least one antibiotic can be administered to alter thebacteria in the patient. As another way of preparing the patient foradministration of the microbial ecosystem, a standard colon-cleansingpreparation can be administered to the patient to substantially emptythe contents of the colon, such as used to prepare a patient for acolonscopy. By “substantially emptying the contents of the colon,” thisapplication means removing at least 75%, at least 80%, at least 90%, atleast 95%, or about 100% of the contents of the ordinary volume of coloncontents. Antibiotic treatment can precede the colon-cleansing protocol.

If a patient has received an antibiotic for treatment of an infection,or if a patient has received an antibiotic as part of a specificpretreatment protocol, in one embodiment, the antibiotic can be stoppedin sufficient time to allow the antibiotic to be substantially reducedin concentration in the gut or vagina before the pharmaceuticalcomposition is administered. In one embodiment, the antibiotic can bediscontinued 1, 2, or 3 days before the administration of thepharmaceutical composition. In another embodiment, the antibiotic can bediscontinued 3, 4, 5, 6, or 7 antibiotic half-lives beforeadministration of the pharmaceutical composition. In another embodiment,the antibiotic can be chosen so the bacterial constituents in thepharmaceutical composition have an MIC50 that is higher than theconcentration of the antibiotic in the gut or vagina.

MIC50 of the bacterial constituents in the pharmaceutical compositioncan be determined by methods well known in the art (see, e.g., Reller etal. (2009) CLINICAL INFECTIOUS DISEASES 49(11):1749-1755). In such anembodiment, the additional time between antibiotic administration andadministration of the pharmaceutical composition is not necessary. Ifthe pretreatment protocol is part of treatment of an acute infection,the antibiotic can be chosen so that the infection is sensitive to theantibiotic, but the bacterial constituents in the pharmaceuticalcomposition are not sensitive to the antibiotic.

Routes of administration. Compositions can be administered by any routesuitable for the delivery of disclosed compositions for treating,inhibiting, or preventing a dysbiosis, or an diseases and disordersassociated with a dysbiosis, include, but are not limited to orally,sublingually, rectally, parentally (e.g., intravenous injection (i.v.).intracranial injection (i.e.); intramuscular injection (i.m.),intraperitoneal injection (i.p.), and subcutaneous injection (s.c.) andintraosseous infusion (i.e.)) transdermally, extracorporeally,inhalation, topically or the like, including topical intranasaladministration or administration by inhalant.

In some embodiments, the subject is fed a meal within one hour ofadministration of the pharmaceutical composition. In another embodiment,the subject is fed a meal concurrently with administration of thepharmaceutical composition.

In some embodiments, the therapeutic composition is administered atintervals greater than two days, such as once every three, four, five orsix days, or every week or less frequently than every week. In otherembodiments, the preparation is administered intermittently according toa set schedule, e.g., once a day, once weekly, or once monthly, or whenthe subject relapses from the primary illness.

In certain embodiments, the pharmaceutical composition is administeredenterically. This preferentially includes oral administration, or by anoral or nasal tube (including nasogastric, nasojejunal, oral gastric, ororal jejunal). In other embodiments, administration includes rectaladministration (including enema, suppository, or colonoscopy). Thepharmaceutical composition can be administered to at least one region ofthe gastrointestinal tract, including the mouth, esophagus, stomach,small intestine, large intestine, and rectum. In some embodiments, it isadministered to all regions of the gastrointestinal tract. Thepharmaceutical compositions can be administered orally in the form ofmedicaments such as powders, capsules, tablets, gels or liquids. Thebacterial compositions can also be administered in gel or liquid form bythe oral route or through a nasogastric tube, or by the rectal route ina gel or liquid form, by enema or instillation through a colonoscope orby a suppository. In certain embodiments of the above invention, themicrobial composition is administered enterically with one or moreprebiotics.

If the composition is administered colonoscopically and, optionally, ifthe composition is administered by other rectal routes (such as an enemaor suppository) or even if the subject has an oral administration, thesubject can have a colon-cleansing preparation. The colon-cleansingpreparation can facilitate proper use of the colonoscope or otheradministration devices, but even when it does not serve a mechanicalpurpose, it can also maximize the proportion of the bacterial cells inthe pharmaceutical composition relative to the other organismspreviously residing in the gastrointestinal tract of the subject. Forexample, the colon cleansing preparation may maximize the amount ofbacterial entities of the bacterial composition that reach and/orengraft in the gastrointestinal tract of the subject.

Dosages and schedule for administration. In some embodiments, thepharmaceutical compositions are provided in a dosage form. In certainembodiments, the dosage form is designed for administration of at leastone OTU or combinations thereof disclosed herein, wherein the totalamount of pharmaceutical composition administered is selected from 0.1ng to 10 g, 10 ng to 1 g, 100 ng to 0.1 g, 0.1 mg to 500 mg, 1 mg to 100mg, or from 10-15 mg. In other embodiments, the pharmaceuticalcomposition is consumed at a rate of from 0.1 ng to log a day, long to 1g a day, 100 ng to 0.1 g a day, 0.1 mg to 500 mg a day, 1 mg to 100 mg aday, or from 10-15 mg a day, or more.

In certain embodiments, the treatment period is at least 1 day, at least2 days, at least 3 days, at least 4 days, at least 5 days, at least 6days, at least 1 week, at least 2 weeks, at least 3 weeks, at least 4weeks, at least 1 month, at least 2 months, at least 3 months, at least4 months, at least 5 months, at least 6 months, or at least 1 year. Insome embodiments the treatment period is from 1 day to 1 week, from 1week to 4 weeks, from 1 month, to 3 months, from 3 months to 6 months,from 6 months to 1 year, or for over a year.

In one embodiment, between about 10⁵ and about 10¹² CFUs total can beadministered to the patient in a given dosage form. In anotherembodiment, an effective amount can be provided in from 1 to 500 ml orfrom 1 to 500 grams of the pharmaceutical composition having from 10⁷ to10¹¹ bacteria per ml or per gram, or, for example, a capsule, tablet orsuppository may contain from 1 mg to 1000 mg lyophilized powder havingfrom 10⁷ to 10¹¹ CFUs. Those receiving acute treatment can receivehigher doses than those who are receiving chronic administration (suchas hospital workers or those admitted into long-term care facilities).

Any of the pharmaceutical compositions described herein can beadministered once on a single occasion or on multiple occasions, such asonce a day for several days or more than once a day on the day ofadministration (including twice daily, three times daily, or up to fivetimes daily). In another embodiment, the preparation can be administeredintermittently according to a set schedule, e.g., once weekly, oncemonthly, or when the patient relapses from the primary illness.

Combination therapy. The pharmaceutical compositions, with or withoutone or more prebiotics, can be administered with other agents in acombination therapy mode, including anti-microbial agents.Administration can be sequential, over a period of hours or days, orsimultaneous.

In one embodiment, the microbial compositions, with or without one ormore prebiotics, are included in combination therapy with one or moreanti-microbial agents, which include anti-bacterial agents, anti-fungalagents, anti-viral agents and anti-parasitic agents.

Anti-bacterial agents can include cephalosporin antibiotics (cephalexin,cefuroxime, cefadroxil, cefazolin, cephalothin, cefaclor, cefamandole,cefoxitin, cefprozil, and ceftobiprole); fluoroquinolone antibiotics(cipro, levaquin, floxin, tequin, avelox, and norflox); tetracyclineantibiotics (tetracycline, minocycline, oxytetracycline, anddoxycycline); penicillin antibiotics (amoxicillin, ampicillin,penicillin V, dicloxacillin, carbenicillin, vancomycin, andmethicillin); and carbapenem antibiotics (ertapenem, doripenem,imipenem/cilastatin, and meropenem).

Anti-viral agents can include Abacavir, Acyclovir, Adefovir, Amprenavir,Atazanavir, Cidofovir, Darunavir, Delavirdine, Didanosine, Docosanol,Efavirenz, Elvitegravir, Emtricitabine, Enfuvirtide, Etravirine,Famciclovir, Foscarnet, Fomivirsen, Ganciclovir, Indinavir, Idoxuridine,Lamivudine, Lopinavir Maraviroc, MK-2048, Nelfinavir, Nevirapine,Penciclovir, Raltegravir, Rilpivirine, Ritonavir, Saquinavir, Stavudine,Tenofovir Trifluridine, Valaciclovir, Valganciclovir, Vidarabine,Ibacitabine, Amantadine, Oseltamivir, Rimantidine, Tipranavir,Zalcitabine, Zanamivir and Zidovudine.

Anti-fungal agents include, but are not limited, to polyene antifungalssuch as natamycin, rimocidin, filipin, nystatin, amphotericin B,candicin, and hamycin; imidazole antifungals such as miconazole,ketoconazole, clotrimazole, econazole, omoconazole, bifonazole,butoconazole, fenticonazole, isoconazole, oxiconazole, sertaconazole,sulconazole, and tioconazole; triazole antifungals such as fluconazole,itraconazole, isavuconazole, ravuconazole, posaconazole, voriconazole,terconazole, and albaconazole; thiazole antifungals such as abafungin;allylamine antifungals such as terbinafine, naftifine, and butenafine;and echinocandin antifungals such as anidulafungin, caspofungin, andmicafungin. Other compounds that have antifungal properties include, butare not limited to polygodial, benzoic acid, ciclopirox, tolnaftate,undecylenic acid, flucytosine or 5-fluorocytosine, griseofulvin, andhaloprogin.

In one embodiment, the pharmaceutical compositions are administered incombination with one or more corticosteroids, mesalazine, mesalamine,sulfasalazine, sulfasalazine derivatives, immunosuppressive drugs,cyclosporin A, mercaptopurine, azathiopurine, prednisone, methotrexate,antihistamines, glucocorticoids, epinephrine, theophylline, cromolynsodium, anti-leukotrienes, anti-cholinergic drugs for rhinitis,anti-cholinergic decongestants, mast-cell stabilizers, monoclonalanti-IgE antibodies, vaccines, and combinations thereof.

The pharmaceutical compositions described herein have beneficial effectsfor the subject locally, at the site of administration (e.g., in thegastrointestinal tract for compositions administered orally, or in thevagina for compositions administered vaginally), as previouslydescribed. Surprisingly, the pharmaceutical compositions describedherein may also be used to correct or prevent a dysbiosis at a sitedistal to the site of administration, intended engraftment, or intendedcolonization of a composition, e.g., a probiotic composition, of theinvention. For example, if a probiotic composition is administeredvaginally, a distal effect of the composition would occur outside thevagina. Similarly, if a probiotic composition is administered to theskin, e.g., through a skin patch, transdermal lotion, etc., a distaleffect of the composition would occur in a niche other than the skin. Ifa probiotic composition is administered to the lungs, e.g., in aninhalable formulation, a distal effect of the composition would occuroutside the lungs. If a probiotic composition is administered to theear, eye, nose, etc., a distal effect of the composition would occur ata site other than the site of administration, engraftment, orcolonization of the composition (i.e., distal to the ear, distal to theeye, distal to the nose, etc.).

Distal sites include but are not limited to the liver, spleen, fallopiantubes and uterus. Other distal sites include skin, blood and lymphnodes. In other embodiments, the distal site is placenta, spleen, liver,uterus, blood, eyes, ears, lungs, liver, pancreas, brain, embryonic sac,or vagina. In another embodiment, the distal site is vagina, skin,lungs, brain, nose, ear, eyes/conjunctiva, mouth, circulatory system,e.g., blood, placenta, reproductive tract, cardiovascular system, and/ornervous system. A probiotic composition may have an effect on themicrobiota of more than one distal site in a subject. For example, insome embodiments, a probiotic composition modulates the microbiota ofone or more sites distal to the site of administration, engraftment, orcolonization, e.g., one or more of placenta, spleen, liver, uterus,blood, eyes, ears, lungs, liver, pancreas, brain, embryonic sac, vagina,skin, brain, nose, mouth, reproductive tract, cardiovascular system,and/or nervous system. In preferred embodiments, the probioticcomposition contains a immunomodulatory bacteria, e.g., aanti-inflammatory bacteria.

Without wishing to be bound by theory, the probiotic compositions of theinvention may impact sites distal sites in several ways.

Pharmaceutical compositions described herein can correct or treat adistal dysbiosis by correcting the imbalance in microbial diversity thatis present at the distal site. Bacteria contained in the pharmaceuticalcomposition can correct the distal dysbiosis directly, by translocatingto the distal site. Bacteria contained in the pharmaceutical compositioncan also correct the distal dysbiosis indirectly, by promotingtranslocation of other gut commensals to the distal site, or bymodifying the microenvironment of the distal site to create conditionsthat restore a healthy microbiome, e.g., by reducing inflammation.

A distal dysbiosis includes disruptions in the normal diversity and/orfunction of the microbial network in a subject at a site other than thegastrointestinal tract, which is generally the site of administration ofprobiotics provided orally. In cases where a probiotic composition isadministered vaginally to a subject, a distal dysbiosis can includedisruptions in the normal diversity and/or function of the microbialnetwork in a subject at a site other than the vagina.

In order to characterize a distal dysbiosis, provided are methods ofdetecting, quantifying and characterizing 16S, 18S and ITS signatures inimmune organs, such as the lymph nodes, spleen, etc. Moreover, providedare methods of detecting bacterial and fungal components typicallyassociated with one microbiota in a distal site, often associating (in aphysiological or pathological manner) with the microbiota of that distalsite. For example, bacteria normally detected in the GI tract or vaginaare detected in distal sites, for example, the blood.

In one embodiment, a bacterial strain present in the pharmaceuticalcomposition engrafts in the gastrointestinal tract of a subject, andtranslocates to a distal site, thereby augmenting the bacterial strainpresent in the pharmaceutical composition at the distal site. In oneembodiment, the bacterial strain present in the pharmaceuticalcomposition is not detectably present at the distal site prior toadministration of the pharmaceutical composition.

In another embodiment, a bacterial strain present in the pharmaceuticalcomposition is augmented in the gastrointestinal tract of a subjectwithout engraftment, and translocates to a distal site, therebyaugmenting the bacterial strain present in the pharmaceuticalcomposition at the distal site. In one embodiment, the bacterial strainpresent in the pharmaceutical composition is not detectably present atthe distal site prior to administration of the pharmaceuticalcomposition.

In another embodiment, a bacterial strain present in the pharmaceuticalcomposition modulates the microenvironment of the gut, augmenting asecond bacterial strain present within the gut microbiota. The secondbacterial strain augmented in the gut translocates to a distal site,thereby augmenting the second bacterial strain at the distal site. Inembodiments, the second bacterial strain is not present in thepharmaceutical composition. In some embodiments, the bacterial strainpresent in the pharmaceutical composition is an immunomodulatorybacteria, e.g., an anti-inflammatory bacteria. Modulation of themicroenvironment of the gut may include, for example, alteration ofcytokines secreted by host cells in and around the gut, reducinginflammation in the gut, increasing secretion of short chain fatty acidsin the gut, or altering the proportion of immune cell subpopulations inthe gut, each of which impacts the gut microbiome. Modulation of themicroenvironment of the gut can include increasing or decreasing overallmicrobial diversity.

In another embodiment, a bacterial strain present in the pharmaceuticalcomposition modulates the microenvironment at a distal site in asubject, thereby augmenting a second bacterial strain at the distalsite. In embodiments, the second bacterial strain is not present in thepharmaceutical composition. In some embodiments, the bacterial strainpresent in the pharmaceutical composition is an immunomodulatorybacteria, e.g., an anti-inflammatory bacteria. Immunomodulatory bacteriacan modulate the microenvironment at a distal site in a subject by, forexample, reducing systemic inflammation. This can be achieved byaltering the profile of cytokine expression by immune cells whichcirculate throughout the body, or altering the proportion of immune cellsubpopulations which circulate throughout the body. Bacterial strainspresent in the pharmaceutical composition can also modulate intestinalpermeability, e.g., by secretion of short chain fatty acids, whichimpacts the microenvironment of distal sites. In addition oralternatively, bacterial strains present in the pharmaceuticalcomposition can increase or decrease overall microbial diversity.

Accordingly, the pharmaceutical compositions described herein mayadditively or synergistically elicit an immunomodulatory response eitherdistally, e.g., in which enteral administration of microbes results inaltering the immune response at a site outside the gastrointestinaltract such as the skin or liver, or locally, e.g. the enteraladministration of microbes results in altering the immune response inthe gastrointestinal tract, e.g., in the intestines.

The immune system of a subject and the microbiome of the subject areclosely linked, and interact systemically. Disruptions to themicrobiome, both in the gastrointestinal tract and at distal sites, canhave profound effects throughout the body of the subject. In particular,disruptions to the microbiome increase systemic inflammation andintestinal barrier dysfunction in a subject. Increased inflammation andintestinal barrier dysfunction negatively impact the health of thesubject in many ways, by contributing to a wide range of inflammatoryand autoimmune conditions distal to the gastrointestinal tract.Conversely, increased inflammation in a subject leads to disruptions inthe subject's microbiome, and disruptions to the microbiome lead in turnto further increases in inflammation. Administration of a pharmaceuticalcomposition containing immunomodulatory bacteria can reduce inflammationin the gastrointestinal tract and restore intestinal barrier integrity,resulting in a reduction in inflammation at sites distal to thegastrointestinal tract, and improvement in the symptoms of autoimmune orinflammatory disorders associated with systemic inflammation.Administration of a pharmaceutical composition containing bacterialstrains that secrete short chain fatty acids are also capable ofreducing inflammation restoring intestinal barrier integrity.

The pharmaceutical compositions and methods described herein can preventor treat the loss or reduction of barrier function recognized to occurduring dysbiosis or in the shift in one or more microbiotal populationsthat give rise to the dysbiosis. The loss of barrier function results insystemic seeding of bacterial populations resulting in dysbioticactivity, and in some events, the loss of barrier function results in alocal reseeding of the bacterial populations. In both situations, theresulting immune activation leads to pathogenic inflammatory and immuneresponses. In response, provided are compositions that are capable ofrestoring barrier function, restoring the normal microbiotal components,and reducing (e.g., suppressing) immune/inflammatory response. In somecompositions, provided are antibiotic agents that remove the existingmicroflora in a target niche, while newly administered or recruitedbacteria populate (or re-populate) the target niche. Co-administrationor co-formulation with a carbohydrate may synergistically affect thispopulation/repopulation technique.

Disorders associated with a dysbiosis, i.e., a gastrointestinaldysbiosis or a distal dysbiosis, which increases systemic inflammationand/or reduces intestinal barrier integrity include, for example,autoimmune or inflammatory disorders, Crohn's Disease, vaginaldysbiosis, and transplant disorders such as graft-versus-host disease.These disorders can be treated by administration (e.g., oraladministration) of pharmaceutical compositions containingimmunomodulatory (e.g., anti-inflammatory) bacterial strains.

The pharmaceutical compositions described herein may additively orsynergistically reduce the number of types of autoimmune disease- orinflammatory disease-associated pathogens or pathobionts eitherdistally—e.g., orally-administered microbes reduce the total microbialburden in an organ not in the gastrointestinal tract, orintravaginally-administered microbes reduce the total microbial burdenin an organ that is not the vagina—or locally, e.g., the intestines orvagina, respectively.

Accordingly, in one aspect, the invention provides a method of reducinginflammation in a subject, comprising administering to the subject aprobiotic composition comprising an isolated, anti-inflammatorybacterial population, such that inflammation in the subject is reduced.A systemic reduction in inflammation can modulate the microbiome ofniches distal to the site of administration, intended engraftment, orintended colonization of the bacterial population. The probioticcomposition can contain an excipient useful for formulation as apharmaceutical composition. In instances where the bacterial populationincludes anaerobic bacteria, the excipient can, in one embodiment,reduce exposure of the bacterial population to oxygen.

In a preferred embodiment, administration of the probiotic compositioncan reduce inflammation at a site distal to the site of administration,engraftment, or colonization, such as, for example, vagina, skin, lungs,brain, nose, ear, eyes/conjunctiva, mouth, circulatory system, e.g.,blood, placenta, embryonic sac, reproductive tract, cardiovascularsystem, and/or nervous system. In one embodiment, administration of theprobiotic composition can reduce inflammation at a site selected fromblood, skin, vagina, liver, spleen, fallopian tubes, uterus, or acombination thereof. In one embodiment, administration of the probioticcomposition modulates the microbiome at a distal site.

The anti-inflammatory bacterial population can induce a decrease insecretion of pro-inflammatory cytokines and/or an increase in secretionof anti-inflammatory cytokines by host cells. The anti-inflammatoryproperties of the bacterial population can be determined by methodsdescribed herein or known in the art, for example, by measuringalterations in cytokine secretion by peripheral blood mononuclear cells(PBMCs) exposed to the bacterial population. Anti-inflammatory bacteriacan be selected for inclusion in the probiotic formulation based onmodulation of particular cytokines of interest. For example,anti-inflammatory bacteria can be selected based on the ability todecrease secretion of one or more pro-inflammatory cytokines, e.g.,IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1, MIP1α, MIP1β, TNFα, andcombinations thereof, and/or the ability to increase secretion of one ormore anti-inflammatory cytokines, e.g., IL-10, IL-13, IL-4, IL-5, TGFβ,and combinations thereof.

In another aspect, the invention provides methods of treating orpreventing a distal dysbiosis in a subject, by administering to thesubject a probiotic composition comprising an isolated bacterialpopulation in an amount sufficient to alter the microbiome at a sitedistal to the site of administration, engraftment, or colonization ofthe bacterial population, such that the distal dysbiosis is treated. Forexample, administration of the probiotic composition may modulate afirst microbiome at the site of administration, engraftment orcolonization of the bacterial population, causing subsequent modulationof a second microbiome at a site that is distinct from the firstmicrobiome, e.g., a distal site.

In one embodiment, the invention provides methods of treating orpreventing a distal dysbiosis, by orally administering a probioticcomposition which alters the microbiome at a site distal to thegastrointestinal tract.

In another aspect, the invention provides a method of treating orpreventing a disorder associated with a distal dysbiosis in a subject inneed thereof, comprising administering to the subject a probioticcomposition comprising an isolated bacterial population in an amountsufficient to alter the microbiome at a site of the distal dysbiosis,such that the disorder associated with the distal dysbiosis is treated.Disorders associated with distal dysbiosis, including disruptions to thesystemic microbiome, are described herein and include, for example,autoimmune or inflammatory disorders such as graft-versus-host disease(GVHD), an inflammatory bowel disease (IBD), ulterative colitis, Crohn'sdisease, multiple sclerosis (MS), systemic lupus erythematosus (SLE),type I diabetes, rheumatoid arthritis, Sjögren's syndrome, and Celiacdisease; transplant disorders such as graft-versus-host disease; andvaginal dysbiosis. In one embodiment, the disorder associated withdistal dysbiosis occurs in the respiratory tract (e.g., lung), includingbut not limited to Cystic Fibrosis and chronic obstructive pulmonarydisorder (COPD).

In one embodiment, the probiotic composition contains a species ofbacteria that is deficient at the site of the distal dysbiosis.Administration of the probiotic composition can increase the quantity ofthe deficient species in the distal microbiome. In one embodiment, thedeficient species is not detectably present at the site of the distaldysbiosis prior to administration of the probiotic composition. In oneembodiment, the species of bacteria in the probiotic compositiontranslocates to the site of the distal dysbiosis.

In another embodiment, the probiotic composition results in augmentationof a species of bacteria not present in the probiotic composition at adistal site. This augmentation can result from, for example,translocation of a species of bacteria not present in the probioticcomposition to the distal site, and/or modulation of themicroenvironment of the distal site in a manner that alters themicrobiome.

In preferred embodiments, the probiotic composition containsimmunomodulatory bacteria, e.g., anti-inflammatory bacteria.

In another aspect, the invention provides a method of reducingintestinal permeability in a subject, by administering a probioticcomposition comprising an isolated bacterial population, whereinadministration of the probiotic composition augments a species ofbacteria that produces short chain fatty acids, such that the intestinalpermeability of the subject is reduced. In other embodiments, intestinalpermeability and disorders associated therewith is improved byadministering a probiotic composition containing mucin-containingbacteria, and/or anti-inflammatory bacteria.

Pharmaceutical compositions useful for correcting or treating a distaldysbiosis, or for treating a disorder distal to the site ofadministration (e.g., the gastrointestinal tract) associated with adysbiosis, can include any of the pharmaceutical compositions describedherein. In exemplary embodiments, a pharmaceutical composition usefulfor correcting or treating a distal dysbiosis includes one or morebacterial strains from Table 1. In other embodiments, the pharmaceuticalcomposition useful for correcting or treating a distal dysbiosisincludes one or more bacterial strains from Table 1A. In otherembodiments, the pharmaceutical composition useful for correcting ortreating a distal dysbiosis includes one or more bacterial strains fromTable 1B. In other embodiments, the pharmaceutical composition usefulfor correcting or treating a distal dysbiosis includes one or morebacterial strains from Table 1C. In other embodiments, thepharmaceutical composition useful for correcting or treating a distaldysbiosis includes one or more bacterial strains from Table 1D. In otherembodiments, the pharmaceutical composition useful for correcting ortreating a distal dysbiosis includes one or more bacterial strains fromTable 1E. In other embodiments, the pharmaceutical composition usefulfor correcting or treating a distal dysbiosis includes one or morebacterial strains from Table 1F. In some embodiments, the pharmaceuticalcomposition contains a single strain of bacteria. In other embodiments,the pharmaceutical composition contains two or more strains of bacteria,e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90,100, 500, 1000 or more strains of bacteria. In other embodiments, thepharmaceutical composition contains or is administered in conjunctionwith a prebiotic, as described herein.

Exemplary pharmaceutical compositions useful for treatment of disordersassociated with a dysbiosis distal to the site of administration (e.g.,the gastrointestinal tract) contain bacterial strains capable ofreducing inflammation in a subject. As described herein, suchimmunomodulatory (anti-inflammatory) bacteria can modulate cytokineexpression by host immune cells, resulting in an overall increase insecretion of anti-inflammatory cytokines and/or an overall decrease insecretion of pro-inflammatory cytokines, systemically reducinginflammation in the subject. In exemplary embodiments, pharmaceuticalcompositions useful for treatment of disorders associated with a distaldysbiosis stimulate secretion of one or more anti-inflammatory cytokinesby host immune cells, such as PBMCs. Anti-inflammatory cytokinesinclude, but are not limited to, IL-10, IL-13, IL-9, IL-4, IL-5, TGFβ,and combinations thereof. In other exemplary embodiments, pharmaceuticalcompositions useful for treatment of disorders associated with a distaldysbiosis inhibit secretion of one or more pro-inflammatory cytokines byhost immune cells, such as PBMCs. Pro-inflammatory cytokines include,but are not limited to, IFNγ, IL-12p70, IL-1α, IL-6, IL-8, MCP1, MIP1α,MIP1β, TNFα, and combinations thereof. Other exemplary cytokines areknown in the art and are described herein. Pharmaceutical compositionscontaining anti-inflammatory bacteria reduce inflammation at the site ofadministration, e.g., in the gastrointestinal tract, as well as atdistal sites throughout the body of the subject.

Other exemplary pharmaceutical compositions useful for treatment ofdisorders associated with a dysbiosis distal to the site ofadministration (e.g., the gastrointestinal tract) contain bacterialstrains capable of altering the proportion of immune subpopulations,e.g., T cell subpopulations, in the subject.

For example, immunomodulatory bacteria can increase or decrease theproportion of Treg cells, Th17 cells, Th1 cells, or Th2 cells in asubject. The increase or decrease in the proportion of immune cellsubpopulations may be systemic, or it may be localized to a site ofaction of the pharmaceutical, e.g., in the gastrointestinal tract or atthe site of a distal dysbiosis. In some embodiments, a pharmaceuticalcomposition comprising immunomodulatory bacteria is used for treatmentof disorders associated with a dysbiosis distal to the site ofadministration (e.g., the gastrointestinal tract) based on the desiredeffect of the pharmaceutical composition on the differentiation and/orexpansion of subpopulations of immune cells in the subject.

In one embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that increase the proportion of Treg cells ina subject. In another embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that decrease the proportion of Treg cells ina subject. In one embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that increase the proportion of Th17 cells ina subject. In another embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that decrease the proportion of Th17 cells ina subject. In one embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that increase the proportion of Th1 cells in asubject. In another embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that decrease the proportion of Th1 cells in asubject. In one embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that increase the proportion of Th2 cells in asubject. In another embodiment, a pharmaceutical composition containsimmunomodulatory bacteria that decrease the proportion of Th2 cells in asubject.

In one embodiment, a pharmaceutical composition containsimmunomodulatory bacteria capable of modulating the proportion of one ormore of Treg cells, Th17 cells, Th1 cells, and combinations thereof in asubject. Certain immune cell profiles may be particularly desirable totreat or prevent particular disorders associated with a dysbiosis. Forexample, treatment or prevention of autoimmune or inflammatory disorderscan be promoted by increasing numbers of Treg cells and Th2 cells, anddecreasing numbers of Th17 cells and Th1 cells. Accordingly,pharmaceutical compositions for the treatment or prevention ofautoimmune or inflammatory disorders may contain pharmaceuticals capableof promoting Treg cells and Th2 cells, and reducing Th17 and Th1 cells.

Distal disorders associated with loss of intestinal barrier function canbe treated or improved by administration of pharmaceutical compositionscontaining bacterial strains that produce short chain fatty acids(SCFAs), such as, for example, butyrate, acetate, propionate, orvalerate, or combinations thereof. Distal disorders associated with lossof intestinal barrier function can be treated or improved byadministration of probiotic compositions containing bacterial strainsthat reduce inflammation, as described herein.

In other embodiments, the distal dysbiosis is caused by a deficiency inmicrobes that produce lactic acid. Accordingly, in one embodiment, theprobiotic composition can contain a species of bacteria that producelactic acid.

EXAMPLES

The invention is further illustrated by the following examples. Theexamples are offered for illustrative purposes only, and are notintended to limit the scope of the present invention in any way. Theentire contents of all references, patents, and published patentapplications cited throughout this application are hereby incorporatedby reference in their entirety.

The practice of the present invention will employ, unless otherwiseindicated, conventional methods of protein chemistry, biochemistry,recombinant DNA techniques and pharmacology, within the skill of theart. Such techniques are explained fully in the literature. See, e.g.,T. E. Creighton, Proteins: Structures and Molecular Properties (W.H.Freeman and Company, 1993); A. L. Lehninger, Biochemistry (WorthPublishers, Inc., current addition); Sambrook, et al., MolecularCloning: A Laboratory Manual (2nd Edition, 1989); Methods In Enzymology(S. Colowick and N. Kaplan eds., Academic Press, Inc.); Remington'sPharmaceutical Sciences, 18th Edition (Easton, Pa.: Mack PublishingCompany, 1990); Carey and Sundberg Advanced Organic Chemistry 3^(rd) Ed.(Plenum Press, Vols A and B, 1992). Enzyme Linked Immunosorbent Assays(ELISAs) and Western blots described below are performed using kitsaccording to the manufacturers' (e.g., Life Technologies, Thermo FisherScientific, New York, USA) instructions.

EXAMPLES

The invention is further illustrated by the following examples. Theexamples are offered for illustrative purposes only, and are notintended to limit the scope of the present invention in any way. Theentire contents of all references, patents, and published patentapplications cited throughout this application are hereby incorporatedby reference in their entirety.

The practice of the present invention will employ, unless otherwiseindicated, conventional methods of protein chemistry, biochemistry,recombinant DNA techniques and pharmacology, within the skill of theart. Such techniques are explained fully in the literature. See, e.g.,T. E. Creighton, Proteins: Structures and Molecular Properties (W.H.Freeman and Company, 1993); A. L. Lehninger, Biochemistry (WorthPublishers, Inc., current addition); Sambrook, et al., MolecularCloning: A Laboratory Manual (2nd Edition, 1989); Methods In Enzymology(S. Colowick and N. Kaplan eds., Academic Press, Inc.); Remington'sPharmaceutical Sciences, 18th Edition (Easton, Pa.: Mack PublishingCompany, 1990); Carey and Sundberg Advanced Organic Chemistry 3^(rd) Ed.(Plenum Press, Vols A and B, 1992). Enzyme Linked Immunosorbent Assays(ELISAs) and Western blots described below are performed using kitsaccording to the manufacturers' (e.g., Life Technologies, Thermo FisherScientific, New York, USA) instructions.

Example 1. Assessment of Intestinal Permeability after Administration ofBacteria, Prebiotic or Combinations Thereof

The main function of the gastrointestinal (GI) tract is to digest andabsorb nutrients from food. The mucosa of the GI tract forms a selectivebarrier between the host and the environment of the gut lumen. Themucosa allows transport of nutrients while restricting passage of largermolecules and bacteria. Impaired barrier integrity is believed tocontribute to the pathogenesis of many disorders including autoimmunediseases, including transplant disorders such asgraft-versus-host-disease (GVHD), and neurological disorders. Disruptionof the intestinal barrier due to toxins, dysbiosis, inflammation orother factors is believed to result in the passage and presentation ofenvironmental antigens to the immune system leading to aberrant immuneresponses. Similarly, the leakage of bacterial endotoxin or other toxicmetabolites into the circulation can lead to systemic inflammationpromoting the development of autoimmunity and neuroinflammation.

Restoration of GI barrier integrity through the administration ofselected prebiotics and/or probiotics represents an approach to correcta basic defect underlying multiple pathological conditions.

In a first set of experiments, intestinal permeability was assessedusing serum endotoxin levels as a marker of gut permeability in micetreated with xylose and/or antibiotics. Basal levels of intestinalpermeability can be measured under disease or normal conditions.Intestinal permeability can be induced in mice through administration ofinflammatory stimuli such as cholera toxin (3 oral gavages of 10 μgcholera toxin, 5 days apart), Poly I:C (3 intraperioneal injections of 1mg/kg, 3 days apart) or dextran sulfate (3% dextran sulfate sodium saltin drinking water for 7 days). Quantitation of intestinal permeabilitywas carried out by quantitatively measuring plasma levels of endotoxinoriginating from gut bacteria using a commercially available chromogenicassay (Lonza, Rockland, Me.). The results of these experiments are shownin FIG. 1 .

Quantitation of intestinal permeability can also be conducted using anumber of alternative methods (reviewed in Bischoff et al, 2014) forexample, by quantifying leakage of fluorescently-labeled high molecularweight dextran (FITC-dextran) into the plasma following oraladministration (oral gavage with 0.6 g/kg 4 kDa FITC-dextran, serumsamples collected 4 hours later and read for fluorescence intensity at521 nm; Hsiao et al, 2013). To study the effect of bacterial strains onintestinal permeability, mice are gavaged orally with 10⁷-10¹⁰ bacterialcells for an average of 5 administrations, typically daily or 2 daysapart. Bacteria can be administered as single strains or combinations ofstrains. The bacteria can be administered alone or in combination with apre-biotic(s). The pre-biotic can be xylose or xylose-containingmolecules as a preferred carbon source for anaerobic bacteria. Otherprebiotics that can be used include, for example, those described inTable 4. After administration of bacteria+/−pre-biotic, intestinalpermability is assessed using the preferred method at the desired timepoint(s) starting on day 1 post-treatment.

As shown in FIG. 1 , C57BL/6 mice were either left untreated or weretreated with xylose at 10 g/L in drinking water from day −7 to day 14;ciprofloxacin (cipro) at 0.25 g/L in drinking water from day −7 to day−2; enrofloxacin (enro) at 0.25 g/L in drinking water from day −7 to day−2; xylose+cipro or xylose+enro. Analysis of serum samples collected ondays 0 and 14 showed that basal levels of serum endotoxin are present innormal mice that remained unchanged in untreated mice. Xylose treatmentreduced these basal levels over time, suggesting an increase in gutbarrier integrity even in normal animals. Antibiotic treatment withcipro, a broad spectrum quinolone antibiotic, or enro, ananaerobe-sparing antibiotic, led to an increase in serum endotoxinlevels (measured 2 days after a 5 day course), likely due to disruptionof the microbiota. Serum endotoxin levels returned to baseline overtime. As shown in FIG. 1 , xylose appeared to counteract the increase inserum endotoxin level caused by cipro, but not enro. The differentialeffect of xylose on these 2 antibiotics may relate to its ability topreserve/promote expansion of anaerobic bacteria, which are killed bycipro but not enro.

Example 2. Immunomodulatory Properties of Different Human CommensalBacteria on Human Peripheral Blood Mononuclear Cells

The microbiota of mammalian hosts is composed of bacterial species thatpossess both pro- and anti-inflammatory properties. In healthyindividuals, a balance or state of eubiosis is maintained that supportsgut barrier integrity, immune containment of commensal bacteria andpromotion of a tolerogenic environment. Under disease conditions,dysbiosis characterized by an imbalance in pro- and anti-inflammatorybacteria results in local inflammation and compromised gut barrierintegrity, leading to systemic inflammation and aberrant immuneresponses. Administration of selected probiotic bacterial strains(+/−prebiotics) that possess anti-inflammatory activity and promoteimmune tolerance represents an approach to correct a basic defectunderlying multiple pathological conditions.

An in vitro system was developed to efficiently test the inflammatoryand immunomodulatory properties of different human commensal bacteria onhuman peripheral blood mononuclear cells (PBMCs). Experiments werecarried out with 21 bacterial candidates to profile theiranti-inflammatory properties against human PBMCs. The innate propertiesof bacteria alone on human PBMCs were tested as well as their ability tocounteract the pro-inflammatory activity of Enterococcus feacalis.

Human PBMCs were isolated from fresh blood by density-gradientcentrifugation using Ficoll (1-4). Freshly isolated PBMCs were plated at1.5×10⁶ cells per ml per well of a 24-well plate in a total volume of 2mls RPMI-1640 medium+5% human serum, and incubated at 37° C./5% CO₂ withthe following:

-   -   (1) 500 μl of different commensal bacteria suspensions at OD 0.8    -   (2) E. faecalis at 10⁷ colony-forming units (cfu)    -   (3) A combination of commensal bacteria (OD 0.8)+E. faecalis        (107 cfu) (4) Complete medium alone as a negative control    -   (5) Bacterial lipopolysaccharide (LPS; 100 ng/ml) as an        immunomodulatory “positive” control        Culture supernatants were collected at 24, 48 and 72 h, and the        cytokine profile was analyzed by Luminex technology according to        manufacturer's instruction (EMD Millipore, Danvers, Mass.).        Cytokine production was detectable in culture supernatants by 24        h with levels increasing over 48-72 h and sometimes exceeding        the range of quantitation. The results are presented in FIGS.        2-5 for all time points. The 24 h time point was chosen as the        optimal time point for further analysis. The 24 h results are        shown as a composite in FIG. 6 and with statistical analysis on        individual cytokines in FIGS. 7-10 . The results represent the        properties of each bacterial species against human PBMCs and        their ability to counteract inflammatory stimulation with E.        faecalis in vitro. It was found that the commensal bacteria        tested have distinct immunomodulatory properties, and most        appear to counteract the inflammatory activity of E. faecalis        for at least one cytokine.

FIG. 2 shows the time course of Th1 related cytokines that were releasedby human PBMCs incubated with Ruminococcus gnavus (Epv 1), Eubacteriumrectale (Epv 2), Blautia luti (Epv 3), Blautia wexlerae (Epv 5) andEnterococcus faecalis (Epv 8), or combinations of each bacterium with E.faecalis. Amounts of Th1-related pro-inflammatory cytokines interferongamma (IFN-γ), interleukin-12 p70 (IL-12p70), interleukin-6 (IL-6),interleukin-2 (IL-2) and tumor necrosis factor alpha (TNFα) released byPBMCs were measured after 24, 48 and 72 hours. As shown in FIG. 2 , allcommensals have unique immunomodulatory properties. As expected, E.faecalis induced high levels of these pro-inflammatory cytokines. Bycomparison, most of the other bacterial candidates induced lower levelsof Th1-related cytokines and were able to counteract the induction ofone or more inflammatory cytokines by E. faecalis. In particular,Blautia luti (Epv 3), showed minimal induction of Th1-related cytokineson its own and was most effective in counteracting induction of thesecytokines by E. faecalis (Epv 8). This profile is desirable for diseaseindications which are primarily driven by Th1 immune responses, such asGVHD.

FIG. 3 shows the time course of Th2 related cytokines that were releasedin cells treated with R. gnavus (Epv 1), E. rectale (Epv 2), B. luti(Epv 3), B. wexlerae (Epv 5) and E. faecalis (Epv 8), or combinationsthereof. Amounts of anti-inflammatory cytokines interleukin-13 (IL-13),interleukin-4 (IL-4) and interleukin-5 (IL-5) released by PBMCs weremeasured after 24, 48 and 72 hours. Each bacterium displayed detectablepattern of cytokine induction and ability to modulate the effect of E.faecalis. Th2-related cytokines are beneficial in counteracting Th1responses. Bacteria capable of promoting Th2 cytokine release aretherefore of interest in Th1-driven diseases. R. gnavus appeared themost active in terms of eliciting Th2 cytokine on its own or in thepresence of E. faecalis.

FIG. 4 shows the time course of Th9, Th17 and Treg cytokines that werereleased in cells treated with R. gnavus (Epv 1), E. rectale (Epv 2), B.luti (Epv 3), B. wexlerae (Epv 5) and E. faecalis (Epv 8), orcombinations thereof. Amounts of interleukin-9 (IL-9), interleukin-17(IL-17) and interleukin-10 (IL-10) released by PBMCs were measured after24, 48 and 72 hours. The activity of IL-9 and IL-17 is context-dependentin that these cytokines can be beneficial under some conditions butdetrimental under other conditions depending on the mechanismsresponsible for disease pathogenesis. For example, IL-17 is expected tocontribute to disease pathogenesis in GVHD but could provide a benefitin Th2-driven disorders. IL-10 produced by regulatory T cells (Treg) isgenerally immunosuppressive and is expected to provide a benefit in mostinflammatory disorders whether Th1- or Th2-driven. As shown in FIG. 4 ,all bacterial candidates elicited IL-9 and IL-17 to varying degrees andB. wexlerae (Epv 5) was the most potent in inducing IL-10.

FIG. 5 shows the time course of monocyte, macrophage andneutrophil-related inflammatory cytokines that were released by PBMCstreated with R. gnavus (Epv 1), E. rectale (Epv 2), B. luti (Epv 3), B.wexlerae (Epv 5) and E. faecalis (Epv 8), or combinations thereof.Amounts of monocyte chemotactic protein 1 (MCP-1), macrophageinflammatory protein 1β (MIP1β), macrophage inflammatory protein 1α(MIP1α), regulated on activation, normal T expressed and secretedprotein (RANTES), interleukin-1α (IL-1α), interleukin-1β (IL1β),interferon α2 (IFN-α2) and interleukin-8 (IL-8) that were released weremeasured after 24, 48 and 72 hours. In general, these cytokinescontribute to inflammation by innate immune effector cells. The bacteriatested showed different degrees of induction and effects on E. faecalis.Overall, E. rectale (Epv 2) and B. luti (Epv 3) were the leastinflammatory and the most effective at countering the effect of E.faecalis (Epv 8).

A composite illustration of the secretion of each of thepro-inflammatory and anti-inflammatory cytokines described above in thepresence of each commensal alone or in combination with EPV8 is graphedrelative to the pro-inflammatory bacterial strain E. faecalis (Epv 8) inFIG. 6 . In the context of GVHD, IFNγ (IFNg), IL-12p70, IL-1α (IL-1α),IL-6, IL-8, MCP1, MIP1α (MIP1α), MIP1β (MIP1b) and TNFα (TNFα) areconsidered pro-inflammatory cytokines. IL-10, IL-13, IL-9, IL-4 and IL-5are considered anti-inflammatory cytokines. IL-17 (IL-17A), IL-9 andIL-2 have context dependent activity. The results are shown as apercentage of Epv 8, where cytokine levels in the presence of E.faecalis after 24 hours is set at 100%. Each commensal has a uniquesignature and each one added alone to human PBMCs appeared to be lessinflammatory than E. fecalis (below 100% for pro-inflammatorycytokines), except for B. wexlerae (Epv 5). When added to PBMCs incombination with E. faecalis, most commensals tested (except for Epv 5)also counteracted the pro-inflammatory activity of E. faecalis (below100% for pro-inflammatory cytokines).

FIGS. 7-10 detail individual cytokine profiles of PBMCs followingexposure to various commensals, alone or in combination with thepro-inflammatory bacterium E. faecalis (Epv8). In particular, FIG. 7shows the effect of R. gnavus (EPV1) on cytokine concentration (pg/ml)either alone or in combination with Epv 8 (E. faecalis).

FIG. 8 shows the effect of E. rectale (EPV 2) on cytokine concentration(pg/ml) either alone or in combination with Epv 8 (E. faecalis). FIG. 9shows the effect of B. luti (EPV 3) on cytokine concentration (pg/ml)either alone or in combination with Epv 8 (E. faecalis). FIG. 10 showsthe effect of B. wexlerae (EPV 5) on cytokine concentration (pg/ml)either alone or in combination with Epv 8 (E. faecalis).

Overall, the foregoing data indicate that, among the bacteria tested,EPV3 has a significantly desirable anti-inflammatory profile for aTh-1-driven condition, such as GVHD while EPV5 has a suboptimalanti-inflammatory profile for GVHD. As shown in FIG. 11 , EPV3 hasrelatively low intrinsic inflammatory activity compared to EPV 8 and isable to reduce the induction of pro-inflammatory cytokines by EPV 8,including IL-6, MCP-1, IL-12p70, and IFNγ which are believed tocontribute to the pathogenesis of GVHD. By comparison, EPV 5 is similarto EPV 8 in terms of induction of pro-inflammatory cytokines and showslittle ability to counteract the induction of pro-inflammatory cytokinesby EPV 8.

Additional bacteria were profiled using this methodology including:Clostridium leptum (EPV 6), Blautia faecis (EPV15), Blautia/Ruminococcusobeum ATCC 29174 (EPV 20), Blautia product ATCC 27340 (EPV 21), Blautiacoccoides ATCC 29236 (EPV 22), Blautia hydrogenotrophica ATCC BAA-2371(EPV-23) and Blautia Hansenii ATCC27752 (EPV 24). Strains freshlyisolated by Epiva from the stool of a normal healthy volunteer were alsoprofiled and included: Eubacterium rectale (EPV 35), a previouslyuncultured Blautia, similar to GQ898099_s S1-5 (EPV 47), a previouslyuncultured Blautia, similar to SJTU_C_14_16 (EPV 51), Blautia wexlerae(SJTU_B_09_77) (EPV 52), Blautia luti ELU0087-T13-S-NI_000247 (EPV 54),Blautia wexlerae WAL 14507 (EPV 64), Blautia obeum (EPV 78),Ruminococcus gnavus (EPV 102) and Blautia luti (BlnIX) (EPV 114).Results focusing on key pro-inflammatory (IL-12p70, IFNγ, IP-10, IL-1RA)and anti-inflammatory (IL-10, IL-4, IL-13) cytokines are shown in FIGS.12-27 . As observed with the initial set of bacterial candidates, eachisolate displayed a defined signature. Candidates for treatment ofautoimmune or inflammatory disorders, such as GVHD, displayed lowinduction of pro-inflammatory cytokines and/or positive induction ofanti-inflammatory cytokines, and had ability to counteract theinflammatory activity of E. faecalis. Bacterial candidates meeting thesecriteria include, for example, EPV 35, 51, 78 and 114.

Taken together, these results show that commensals have distinctimmunomodulatory properties and display a definable signature in termsof their ability to induce cytokines in human host cells, or counteractthe pro-inflammatory activity of another bacterium (E. faecalis).Accordingly, bacterial compositions may be selected in order to achievea desired modulation of pro- and anti-inflammatory cytokines. Forexample, anti-inflammatory bacterial strains may be selected based ontheir ability to reduce key pro-inflammatory cytokines such asinterferon gamma, IL-12p70, IP-10 and IL-1RA and/or increaseanti-inflammatory cytokines such as IL-13, IL-10 and IL-4.

Example 3. Effect of Commensal Human Bacteria on T-Cell Polarization

In order to determine whether exposure to commensal bacteria maypolarize T cells toward a particular phenotype, flow cytometry analysiswas performed on human PBMCs cultured with various commensal bacteria asdescribed above. The cells recovered from culture were washed inphosphate-buffered saline and stained with a cocktail of fluorescentlylabeled antibodies against specific cell surface protein markers toallow for the detection of Th1 cells (CXCR3⁺CCR6⁻), Th2 cells(CXCR3⁻CCR6⁻), Th17 cells (CXCR3⁻CCR6⁺) and Tregs (CD25⁺CD127^(lo)).Negative control wells contained PBMCs in culture medium alone andpositive control wells contained PBMCs+LPS (100 ng/ml) as a known immunestimulus. The commensal bacteria examined included: Epv 1: R. gnavus;Epv 3: B. luti; Epv 2: E. rectale; Epv 5: B. wexlerae; Epv. 8: E.faecalis; Epv 20: B. obeum, ATCC 29174; Epv 21: B. product, ATCC 27340;Epv 24: B. hansenii, ATCC 27752. As shown in FIG. 28 , exposure of humanPBMCs to bacteria did result in a shift in the relative proportion of Tcell populations compared to the PBMCs alone (control) althoughstatistical significance was not achieved in every case. Overall, mostbacteria tested caused an increase in the proportion of T cells with aregulatory phenotype (Tregs) with EPV 21 and EPV 24 having the greatestimpact and EPV8 (E. faecalis) causing little or no increase in Tregs.Most bacteria also caused a decrease in the proportion of Th17 cells, anincrease in Th2 cells and had little or no effect on the proportion ofTh1 cells. This type of analysis indicates that commensal bacteria canmodulate the proportions of effector T cell types and can be used toselect the desired phenotype for a given disease application. Forexample, the optimal T cell profile to address pro-inflammatorydisorders such as GVHD would consist of ↑Treg, ↓Th17, 1 or ↓ unchangedTh1, and ↑Th2. This phenotype was induced by many of the bacteriatested.

Example 4. Pattern of Carbon Source Utilization by Commensal Bacteria

Modulation of the microbiota to correct a dysbiosis associated withpathological conditions can potentially be achieved throughadministration of bacteria (or bacterial combinations) and prebiotic(s)as a carbon source to promote endogenous expansion of beneficialbacteria. Alternatively, prebiotics can be administered in combinationwith bacteria to promote their growth or create a favorable environmentfor their growth. Profiling of carbon source usage by bacterial isolatescan be used to customize and optimize selection of prebiotics forparticular bacterial strains. Profiling of carbon source usage wasconducted on 21 anaerobic commensal bacteria (Table 3) using 96 wellplates from Biolog (Hayward, Calif.) where each well contains adifferent carbon source for a total of 192 different carbon sources(PM01 and PM02A plates). The carbon sources tested are listed in Table4. The assay was conducted according to manufacturer's instructions.Briefly, pre-cultured bacteria were suspended in Biolog assay medium ata 750 nm optical density (OD) of 0.3-0.4 and 100 μl of the suspensionwas transferred to each well of the 96 well PM01 and PM02 assay plates.The plates were then incubated at 37° C. in an anaerobic chamber for 24hr or longer. The amount of growth on each carbon source was evaluatedby measuring the optical density (OD) of each well at 750 nm. Theresults are summarized in FIG. 29 , and indicate that each individualstrain displays a unique pattern of carbon source usage. Interestingly,different isolates of the same species (e.g. B. luti and B. wexlerae)show related (albeit distinct) patterns. Overall, these results indicatethat characterization of carbon source usage for profiling of bacterialcandidates allows optimal selection of prebiotics. Preferred prebioticscan be selected which increase the growth (indicated by an increase inoptical density) of bacterial species contained in probioticcompositions.

Example 5. Normal Human Volunteer Study of a Prebiotic FormulationContaining Xylose

D-xylose is a carbon source generally preferred by anaerobic bacteria.Preliminary results in the mouse indicate that it may act to promote gutbarrier integrity (FIG. 1 ). It is also used as a carbon source byseveral bacterial strains (FIG. 29 ) that were determined to possess adesirable immunological profile for target indications such as GVHD(FIG. 19, 25, 27 ). A parallel, double-blind, 5 cohort escalation foodsafety study was conducted to examine D-xylose in normal humanvolunteers. The study was a double-blind, single-center, parallel groupstudy designed to evaluate the tolerability and potential microbiomechanges induced by ingestion of D-xylose at 5 different amounts inhealthy, adult volunteers enrolled at 1 study center in the UnitedStates (US).

Subjects were screened for eligibility within 21 days prior to the firstplanned ingestion of study sweetener on Day 1 (Baseline). Within each of5 cohorts, eligible subjects were randomly assigned in a double-blinded,6:2 ratio to ingest either D-xylose or the GRAS sweetener Splenda®(control), dissolved into 2 to 6 oz of sterile water and ingested TIDwith meals for a total of 82 ingestions taken over 28 consecutive days.D-xylose ingestion amounts ranged from 1 to 15 g TID (total daily amountof 3 to 45 g), and all subjects randomized to Splenda® ingested 1dissolved, commercially available packet TID (3 packets total per day).Subjects returned to the study center weekly on Days 8, 15, 22, and 28for ingestion, tolerability, and compliance evaluations. Safety wasevaluated on a continual basis through adverse events (AE) monitoring,clinical laboratory measurements, vital sign monitoring, physicalexaminations, electrocardiograms (ECGs), telephone follow-up, andelectronic subject ingestion diaries. Stool was collected pre-ingestionand at pre-specified time points, and post-ingestion samples wereevaluated for changes in the gut microbiome compared with Baseline forall subjects. For subjects who consented to further sampling, additionalstool specimens were used to potentially isolate living bacteria thatcould be categorized for research and potential commercializationpurposes. Serum and urine were collected for measurement of D-xyloselevels and pharmacokinetic (PK) assessments and PK/pharmacodynamics (PD)correlations. Telephone follow-up was conducted as needed, but minimallyonce per week. The total duration for each participant was up to 60days, including the Screening period (Day −21 to 0), the ingestionperiod (Day 1 to 28), and an End-of-Study (EOS) follow-up visitconducted 7 (±3) days after the last ingestion of study sweetener.

Criteria for Evaluation

Safety

Safety was evaluated on a continual basis through AE monitoring,clinical laboratory measurements, vital sign monitoring, physicalexaminations, ECGs, telephone follow-up, and electronic subjectingestion diaries.

Immunology and Other Assessments

Stool was collected at pre-specified pre- and post-ingestion time pointsand post-ingestion samples were evaluated for changes in the gutmicrobiome compared with Baseline. Additional optional specimens werecollected to potentially isolate living bacteria that could becategorized for research and potential commercialization purposes.

Blood was collected at pre-specified pre- and post-ingestion time pointsto evaluate C-reactive protein (CRP), serum cytokines (tumor necrosisfactor alpha [TNF-α], interleukin [IL]-2, IL-6, interferon gamma[IFN-γ], and IL-10), and T-cell markers CD3, CD4, CD8, CD25, and FOXP3.Plasma was also stored and may be tested for biomarkers and/or metabolicmarkers for up to 7 years.

Pharmacokinetics

Blood and urine were collected at pre-specified pre- and post-ingestiontime points to measure D-xylose levels and to characterize the systemicabsorption profiles of D-xylose.

Statistical Methods

Statistical analyses were conducted using SAS®, Version 9.2 (SASInstitute, Inc., Cary, N.C., USA). The sample size calculations wereempiric and based on an estimation of normal healthy volunteervariability in reported symptoms and side effects and not on astatistical method. A weighted randomization scheme was implemented suchthat more subjects were enrolled at the higher D-xylose ingestionamounts to account for potential toxicity-related effects that couldhave resulted in withdrawal and/or analysis ineligibility, and to enablecollection of more data at ingestion amounts for which limited data wereavailable.

Analysis Populations

The safety population comprised all subjects who ingested any amount ofstudy sweetener.

Safety

AEs were coded using the Medical Dictionary for Regulatory Activities(MedDRA), Version 18.0 (Northrup Grumman Corporation, Chantilly, Va.,USA), and summarized by cohort. Laboratory, vital sign, and physicalexamination data were summarized by cohort using descriptive statisticsover time, including statistics for changes from Baseline. ECG findingswere also summarized by cohort over time as well as using frequencycounts and percentages, as normal or abnormal, with the relevance ofabnormalities categorized by clinical significance.

Immunology and Other Assessments

Stool sample compliance was summarized by cohort, using the followingcalculation for each subject:

${{Percentage}\mspace{14mu}{compliance}} = {\frac{{Total}\mspace{14mu}{number}\mspace{14mu}{of}\mspace{14mu}{stool}\mspace{14mu}{samples}\mspace{14mu}{collected}}{{Total}\mspace{14mu}{number}\mspace{14mu}{of}\mspace{14mu}{stool}\mspace{14mu}{samples}\mspace{14mu}{expected}} \times 100}$

A total of 7 stool samples were expected to be collected for eachsubject. Evaluation of changes in the gut microbiome were evaluated instool samples through taxonomic classification, relative and statisticaldifferential abundance analyses by cohort and time point, an alphadiversity analysis calculated using the Shannon diversity index bycohort and time point, a beta diversity analysis using Bray-Curtisdissimilarity and Unifrac distance by subject and time point, and aprincipal coordinates analysis using the beta diversity data.

Summary statistics (n, mean, standard deviation, median, minimum, andmaximum) were presented for serum concentrations of CRP, flow cytometryT-cell markers (CD3, CD4, CD8, CD25, and FOXP3), and cytokines (TNF-α,IL-2, IL-6, IFN-γ, and IL-10) as per their nominal time points.

Pharmacokinetics

Phoenix® WinNonLin®, Version 6.2.1, was used for PK analyses.

Serum D-xylose concentrations were summarized by cohort using nominalsample times according to actual amount received using summarystatistics (n, coefficient of variation [CV], mean, standard deviation[SD], median, minimum, and maximum). Evidence for the occurrence ofsteady-state was assessed graphically by comparing the time course ofeither trough or 2-hour post-ingestion serum concentrations of D-xyloseas different levels of D-xylose. Accumulation was assessed by comparingthe 2-hour post-first-ingestion serum levels with those observed at Week2 (Day 15) and Week 4 (Day 28).

The total amount of D-xylose excreted in urine was analyzed for allsubjects over 5 hours post-ingestion and pooled for analysis; thepooling for analysis reflected the subject mean within a given time ofcollection (e.g., Day 15 and then Day 28) sorted by ingested amount.Urine PK parameters for D-xylose levels included Ae_((0-t)) (cumulativeamount of sweetener recovered in urine) and percent sweetener amountexcreted over a 5-hour period.

Summary of Results

Forty-eight subjects were randomized to ingest either 1 packet ofcommercially-available Splenda® TID (n=12) or D-xylose TID at thefollowing ingestion amounts (n=36 total):

1 g: 6 subjects

2 g: 6 subjects

8 g: 7 subjects

12.5 g: 8 subjects

15 g: 9 subjects

Over the 28-day ingestion period, study sweetener ingestion compliancewas >90% for all subjects. Two subjects (4.2%) discontinued from thestudy prematurely; primary reasons for discontinuation were a protocolviolation (positive urine drug screen) and withdrawal of consent. Theproportion of males (47.9%) and females (52.1%) was balanced, and themajority of subjects were White (89.6%) and not Hispanic or Latino(77.1%). Subject ages spanned a wide range, with a median of 38.3 (range22.5 to 60.5) years for the combined D-xylose cohorts and 43.6 (range24.9 to 64.3) years for the Splenda® cohort.

Safety

D-xylose and Splenda® were both well tolerated, with no new safetyconcerns identified. One subject required a D-xylose reduction from 15 gto 12.5 g TID at the Week 1 (Day 8) visit due to AEs of moderateabdominal distension, diarrhea, and GI pain; no other modifications tosweetener ingestion amounts were implemented.

Overall, 17 subjects (35.4%) experienced at least 1 AE, including ahigher proportion of subjects who ingested any amount of D-xylose (14subjects [38.9%]) than Splenda® (3 subjects [25.0%]). Reported AE ratesincreased with increasing D-xylose ingestion amounts, with incidencesranging from 16.7% in subjects who ingested the 2 lowest amounts (1 and2 g TID) to 66.7% in subjects who ingested the highest amount (15 gTID). AEs reported for more than 1 subject in the D-xylose cohortsincluded diarrhea (3 subjects [8.3%]) and flatulence and GI pain (2subjects [5.6%] each). AEs in the Splenda® cohort included abdominaldistension, flatulence, increased blood creatinine, infrequent bowelmovements, and rhinitis. The incidence of AEs was highest during Weeks 1and 2 (Days 2 through 15), regardless of sweetener type or ingestionamount. During this 2-week period, 18 subjects overall (37.5%)experienced AEs, compared with 7 subjects (14.6%) overall whoexperienced AEs either on Day 1 or after Week 2.

All AEs were mild in severity with the exception of moderate AEsreported for 4 subjects (11.1%) in the D-xylose cohorts. These moderateAEs included abdominal distension, concussion/post-concussion syndrome,diarrhea, GI pain, increased blood bilirubin, and neutropenia.

No SAEs, severe AEs, or subject deaths were reported. One subject in the8 g TID D-xylose cohort experienced non-serious, moderate AEs ofconcussion and post-concussion syndrome that were noted to havecontributed to study discontinuation; however, this subject's primaryreason for discontinuation was withdrawal of consent.

GI-related AEs, which were of special interest, were reported for 7subjects (19.4%) in the D-xylose cohorts and 2 subjects (16.7%) in theSplenda® cohort. GI-related events were mild for all but 1 subject inthe 15 g TID D-xylose cohort who experienced moderate GI-related AEs ofabdominal distension, diarrhea, and GI pain that required reduction ofthe D-xylose ingestion amount to 12.5 g TID.

Eleven subjects (22.9%) experienced at least 1 AE that was considered bythe Investigator to be related to study sweetener, including 9 subjects(25.0%) in the D-xylose cohorts and 2 subjects (16.7%) in the Splenda®cohort. The incidence of sweetener-related AEs appeared to increase withincreasing D-xylose ingestion amounts. Sweetener-related AEs reportedfor more than 1 subject in the D-xylose cohorts included diarrhea (3subjects [8.3%]) and flatulence and GI pain (2 subjects [5.6%] each).Sweetener-related AEs reported in the Splenda® cohort were abdominaldistension, flatulence, and infrequent bowel movements.

No fluctuations in clinical laboratory measurements over time wereconsidered to be clinically meaningful. Categorical shifts from Baselinethat occurred in >10% of subjects in either the combined D-xylose orSplenda® cohorts included decreased or increased glucose (27.7% D-xyloseand 16.7% Splenda®) and decreased absolute neutrophil count (ANC) (13.9%and 8.3%); these shifts were not associated with sweetener type oringestion amount.

Immunology and Other Assessments

To assess the effect of D-xylose on the gut microbiome, this studyincorporated an analysis of alpha diversity, beta diversity, anddifferentially abundant taxa. These factors were assessed both acrosscohorts and over time. Regardless of sweetener ingestion amount, noapparent significant impact on the intra-sample alpha diversity of thegut microbiome was observed, and no significant changes in communitycomposition were observed over time on study. Numerous taxa wereidentified as differentially abundant, but these findings may reflectthe relatively small sample sizes in each cohort.

Across all D-xylose cohorts, 8.3% of subjects with normal serum CRP atBaseline experienced at least 1 post-ingestion CRP value >2.9 mg/L. Asubstantially higher proportion of subjects in the Splenda® cohort(41.7%) had normal serum CRP at Baseline and experienced at least 1post-ingestion CRP value >2.9 mg/L. None of the post-ingestion CRPvalues for any subject were deemed clinically significant.

Because most individual cytokine data points were below the limit ofquantitation (BLQ) and therefore set to zero, cytokine summarystatistics were limited and did not indicate any consistent orclinically meaningful changes over time for either sweetener or anyD-xylose ingestion amount. There was a trend for reduced levels of seruminterferon gamma over time in the 2 g and 15 g D-xylose cohorts (FIG. 30). No consistent or clinically meaningful changes over time in totalT-cells or any T-cell subsets were observed for either sweetener or anyD-xylose ingestion amount.

Pharmacokinetics

Serum D-xylose concentrations increased linearly with increasingingestion amounts. Little to no accumulation of serum D-xylose occurredat Day 15 following 1 g to 12.5 g TID ingestion, while an approximately1.9-fold accumulation ratio was observed in the 15 mg TID cohort(although variability was high). On Day 28, the accumulation ratioranged from 1.08 to 1.31 following 1 g to 12.5 g TID ingestion and 1.68following 15 g TID ingestion, although variability was moderate to highin all but the 8 g TID cohort.

In the 1 g TID cohort, approximately 40% of the ingested amount ofD-xylose was recovered in urine within 5 hours post-ingestion on Days 1,15, and 28. In the 2 g through 15 g TID cohorts, between 23% and 32% ofthe ingested amount of D-xylose was recovered in urine within 5 hourspost-ingestion on Days 1, 15, and 28. The fraction excreted in urine wassimilar among Days 1, 15, and 28.

A review of the time course of serum D-xylose concentrations and thecorresponding urinary excretion profiles indicated high ingestioncompliance.

Changes in the Gut Microbiome

A total of 344 stool samples were collected in OMNIgene-GUT collectionkits and shipped to the GenoFIND laboratory for DNA extraction and V3-V416S amplicon sequencing. There were no major shifts in the microbiomealpha diversity between the different treatment groups (absolute numberof OTUs, abundance of OTUs) or over time on study. There was an overalldecrease in the Chao diversity index over time (indicator of communityrichness −# of singleton, doubleton OTUs), as shown in FIG. 31 .Numerous taxa were identified as differentially abundant, but thisfinding may be attributable to the relatively small sample sizes of eachcohort. Similar observations were made in the mouse study, e.g., xylosetreatment did not cause major shifts in the gut microbiome but showedsome differences at the family level. Overall, these results suggestthat, under the conditions tested in normal individuals and normal mice,ingestion of xylose exerts subtle changes in the gut microbiome. Theimpact of xylose on the microbiome under disease conditions remains tobe determined.

Taken together, the results of this trial show that D-xylose is safe andwell-tolerated, and indicate that prebiotic formulations containingxylose may reduce inflammation in a subject, resulting in reduction ofserum levels of pro-inflammatory cytokines.

Example 6. Distal Augmentation

The trillions of organisms forming the microbiome function as an organsystem interconnected throughout the body. The possibility thatmodification of the microbiome in a given physical location mayinfluence the microbiome at other sites in the body (distalaugmentation) was investigated. Seven week old C57Bl/6 female mice wereacclimatized for 7 days prior to the start of the study by dailyhandling and shuffling between cages. All mice were housed at three miceper cage in individually vented cages (Thoren, Hazleton, Pa.). At day 0,baseline fresh fecal pellets, and vaginal lavages with 100 μL of steriledouble-distilled water were collected and immediately frozen at −80° C.for microbiome analysis. After baseline collection, mice were given todrink either autoclaved water (N=6) or 0.5 mg/L of the antibioticvancomycin in autoclaved water (N=6) ad libitum. Water alone is notexpected to influence the microbiome and acted as a negative control.Oral vancomycin is poorly absorbed from the gut and its ingestion doesnot result in significant levels of drug in the body (Rao et al, 2011).The impact of oral vancomycin is therefore expected to be limited to thegastrointestinal tract such that microbiome changes elsewhere in thebody (e.g. vagina) would be attributable to distal augmentation. At day6, fresh fecal pellets and vaginal lavages with 100 μL of steriledouble-distilled water were collected and immediately stored at −80° C.for microbiome analysis.

Isolation and sequencing of microbial DNA from the stool and vaginalsamples was performed by DNA Genotek (Ottawa, ON, Canada). The V3-V4region of the 16S ribosomal subunit was amplified with custom PCRprimers and sequenced on an Illumina MiSeq to a minimum acceptable readdepth of 25,000 sequences per sample. The widely accepted read depthrequirement for accurate taxonomic profiling is 15,000-100,000 reads(Illumina, 2014). A closed-reference taxonomic classification wasperformed, where each sequence was aligned to the SILVA referencedatabase, version 123. Sequences were aligned using the UCLUST algorithmincluded in QIIME version 1.9.1 (Caporaso et al., 2010). A minimumthreshold of 97% sequence identity was used to classify sequencesaccording to representative sequences in the database. At 97% sequenceidentity, each OTU represents a genetically unique group of biologicalorganisms. These OTU's were then assigned a curated taxonomic labelbased on the seven level SILVA taxonomy.

As expected, oral vancomycin treatment had a strong impact on themicrobiome of the gut. As shown by principal component analysis (PCA) atthe family level, the day 0 to day 6 pattern in fecal samples wasclearly different in the control vs oral vancomycin group (FIG. 32 ).Interestingly, the day 0 to day 6 pattern in the vaginal samples alsoshowed an overall difference between the PBS and oral vancomycin groupseven though the vaginal environment is not exposed to vancomycinfollowing oral administration of the antibiotic (FIG. 32 ). In addition,some bacterial species were detected at low frequency in vaginal samplesof the vancomycin-treated group at day 6 (median abundance ofapproximately 0.00002%) that were not present at day 0 (Table 5). Theseresults support the concept of distal augmentation whereby modificationof the microbiome at one site also has an impact at a distal site(s).This finding opens the possibility of modulating the microbiome, forexample at the level of the gut, to effect therapeutic changes in themicrobiome at other sites, for example the lung.

Example 7. Co-Culture of Bacteria Plus Prebiotic and Host-Cells andAnalysis of Host Cell Cytokine Response

The following work is done in the presence and absence (as a control) ofone or more selected prebiotic carbohydrates. This assay may be used totest or confirm the ability of a prebiotic-bacterium pair to elicit animmunomodulatory response such that the production or release ofproinflammatory cytokines decreases and/or the production or release ofanti-inflammatory cytokines increases, may be used to evaluate thedifference in cytokine response in the presence or absence of aprebiotic mixture, and/or may be used to evaluate an array of prebioticcandidates. Clostridales bacteria are obtained from the ATCC or purifiedfrom a human donor and cultured in brain-heart infusion broth at 37° C.The bacteria are harvested by centrifugation (3000 g, 15 minutes) after24 hours of stationary growth. To test the effects of spores on humanintestinal cells and/or human peripheral blood mononuclear cells(huPBMC), bacteria are first heat killed (95° C., 30 minutes) before thecentrifugation step. Bacteria (or spores) are washed three times with1×PBS (pH 7.2, Gibco BRL) and subsequently diluted to obtain final celldensities of 10⁶ and 10⁷ colony forming units (cfu)/ml in RPMI 1640medium (Gibco BRL).

Human enterocyte-like CaCO-2 cells (passage 60-65) are seeded at adensity of 2.5×10⁵ cells/ml on 25 mm cell culture inserts (0.4 mnucleopore size; Becton Dickinson). The inserts are placed into six welltissue culture plates (Nunc) and cultured 18-22 days at 37° C./10% CO₂in DMEM (glutamine, high glucose; Amimed) supplemented with 20%decomplemented fetal calf serum (56° C., 30 minutes; Amimed), 1% MEMnon-essential amino acids (Gibco BRL), 10 μg/ml gentamycin (Gibco BRL),and 0.1% penicillin/streptomycin (10 000 IU/ml/10 000 UG/ml; Gibco BRL).The cell culture medium is changed every second day until the cells arefully differentiated. Transepithelial electrical resistance (TEER) isdetermined continuously in confluent CaCO-2 monolayers using aMultiCell-ERS voltmeter/ohmmeter or as described in Example 44.

Tissue culture inserts covered with CaCO-2 cell monolayers are washedtwice with prewarmed RPMI 1640 medium and transferred to six well tissueculture plates. 2 mL culture medium is added to the apical andbasolateral compartments of the transwell cell culture system.

Next, the apical surface of CaCO-2 monolayers is challenged by additionof 10⁶ or 107 cfu/ml of Clostridiales bacteria or spores, in the absenceof gentamicin. After four hours, gentamicin is added (at 150 pg/mL) tostop bacterial growth and metabolite secretion. CaCO-2 cells arestimulated with the bacteria or spores for 6-36 hours in a 37° C., 10%CO₂ incubator. Then the CaCO-2 cells are collected, washed once withcold 1×PBS (pH 7.2), and lysed in denaturation solution for RNAextraction (Micro RNA Isolation Kit, Stratagene). Cellular lysates arestored at −20° C. and cell culture supernatants are collected from theapical compartment and frozen at −20° C. The immune response of CaCO-2cells is monitored by analysis of cytokine gene transcription (TNF-α,IL-8, monocyte chemoattracting protein 1 (MCP-1), TGF-β, IL-12, IFN-γ,IL-4, IL-10) using a reverse transcription-polymerase chain reaction(RT-PCR) technique and determination of cytokine secretion in cellculture supernatants using an ELISA (Haller D, Bode C, Hammes W P,Pfeifer A M A, Schiffrin E J, Blum S, 2000. Non-pathogenic bacteriaelicit a differential cytokine response by intestinal epithelialcell/leucocyte co-cultures. Gut. 47:79-97).

REFERENCES

-   1. Bischoff, S C, Giovanni, B, Buuman, W, Ockhuizen, T, Schulzke,    J-D, Serino, M, Tilg, H, Watson, A and Wells, J M. (2014) Intestinal    permeability—a new target for disease prevention and therapy. BMC    GASTROENTEROLOGY 14: 189.-   2. Boyum, A. (1968) Isolation of mononuclear cells and granulocytes    from human blood. Scand. J. CLIN. LAB. INVEST. 21, Suppl 97 (Paper    IV), 77-89.-   3. Boyum A. (1976) Isolation of lymphocytes, granulocytes and    macrophages. SCAND J IMMUNOL. (Suppl 5):9-15.-   4. Bach M K, Brashler J R. (1970) Isolation of subpopulations of    lymphocytic cells by the use of isotonically balanced solutions of    Ficoll. I. Development of methods and demonstration of the existence    of a large but finite number of subpopulations. EXP CELL RES.    61:387-96.-   5. Fotino, M., Merson, E. J. and Allen, F. H. (1971) Micromethod for    rapid separation of lymphocytes from peripheral blood. ANN. CLIN.    LAB. SCI. 1:131-133.-   6. Hsiao, E Y, McBride, S W, Hsien, S, Sharon G, Hyde, E R, McCue T,    Codelli, J A, Chow, J, Reisman, S E, Petrosino, J F, Patterson, P H    and Mazmanian, S K (2013) Microbiota modulate behavioral and    physiological abnormalities associated with neurodevelopmental    disorders. CELL 155: 1451-1463.-   7. Caporaso, J. G., Kuczynski, J., Stombaugh, J., Bittinger, K.,    Bushman, F. D., Costello, E. K., Knight, R. (2010). QIIME allows    analysis of high-throughput community sequencing data. NATURE    METHODS 7 (5): 335-336. doi:10.1038/nmeth.f.303-   8. Illumina. (2014). Frequently Asked Questions: 16S Metagenomic    Sequencing. Retrieved from    http://www.illumina.com/content/dam/illuminamarketing/documents/products/other/16smetagen    omics-faq-1270-2014-003.pdf-   9. Rao S, Kupfer Y, Pagala M, Chapnick E and Tessler S. (2011)    Systemic absorption of oral vancomycin in patients with Clostridium    difficile infection. SCAND J INFECT DIS 5: 386-388.

TABLE 1 See, e.g., WO 2014/121304 SEQ ID Public DB Spore Pathogen OTUNumber Accession Clade Former Status Eubacterium saburreum 858 AB525414clade_178 Y N Eubacterium sp. oral clone IR009 866 AY349376 clade_178 YN Lachnospiraceae bacterium ICM62 1061 HQ616401 clade_178 Y NLachnospiraceae bacterium MSX33 1062 HQ616384 clade_178 Y NLachnospiraceae bacterium oral taxon 107 1063 ADDS01000069 clade_178 Y NAlicyclobacillus acidocaldarius 122 NR_074721 clade_179 Y N Clostridiumbaratii 555 NR_029229 clade_223 Y N Clostridium colicanis 576 FJ957863clade_223 Y N Clostridium paraputrificum 611 AB536771 clade_223 Y NClostridium sardiniense 621 NR_041006 clade_223 Y N Eubacterium budayi837 NR_024682 clade_223 Y N Eubacterium moniliforme 851 HF558373clade_223 Y N Eubacterium multiforme 852 NR_024683 clade_223 Y NEubacterium nitritogenes 853 NR_024684 clade_223 Y N Anoxybacillusflavithermus 173 NR_074667 clade_238 Y N Bacillus aerophilus 196NR_042339 clade_238 Y N Bacillus aestuarii 197 GQ980243 clade_238 Y NBacillus amyloliquefaciens 199 NR_075005 clade_238 Y N Bacillusanthracis 200 AAEN01000020 clade_238 Y Category-A Bacillus atrophaeus201 NR_075016 clade_238 Y OP Bacillus badius 202 NR_036893 clade_238 YOP Bacillus cereus 203 ABDJ01000015 clade_238 Y OP Bacillus circulans204 AB271747 clade_238 Y OP Bacillus firmus 207 NR_025842 clade_238 Y OPBacillus flexus 208 NR_024691 clade_238 Y OP Bacillus fordii 209NR_025786 clade_238 Y OP Bacillus halmapalus 211 NR_026144 clade_238 YOP Bacillus herbersteinensis 213 NR_042286 clade_238 Y OP Bacillusidriensis 215 NR_043268 clade_238 Y OP Bacillus lentus 216 NR_040792clade_238 Y OP Bacillus licheniformis 217 NC_006270 clade_238 Y OPBacillus megaterium 218 GU252124 clade_238 Y OP Bacillus nealsonii 219NR_044546 clade_238 Y OP Bacillus niabensis 220 NR_043334 clade_238 Y OPBacillus niacini 221 NR_024695 clade_238 Y OP Bacillus pocheonensis 222NR_041377 clade_238 Y OP Bacillus pumilus 223 NR_074977 clade_238 Y OPBacillus safensis 224 JQ624766 clade_238 Y OP Bacillus simplex 225NR_042136 clade_238 Y OP Bacillus sonorensis 226 NR_025130 clade_238 YOP Bacillus sp. 10403023 MM10403188 227 CAET01000089 clade_238 Y OPBacillus sp. 2_A_57_CT2 230 ACWD01000095 clade_238 Y OP Bacillus sp.2008724126 228 GU252108 clade_238 Y OP Bacillus sp. 2008724139 229GU252111 clade_238 Y OP Bacillus sp. 7_16AIA 231 FN397518 clade_238 Y OPBacillus sp. AP8 233 JX101689 clade_238 Y OP Bacillus sp. B27(2008) 234EU362173 clade_238 Y OP Bacillus sp. BT1B_CT2 235 ACWC01000034 clade_238Y OP Bacillus sp. GB1.1 236 FJ897765 clade_238 Y OP Bacillus sp. GB9 237FJ897766 clade_238 Y OP Bacillus sp. HU19.1 238 FJ897769 clade_238 Y OPBacillus sp. HU29 239 FJ897771 clade_238 Y OP Bacillus sp. HU33.1 240FJ897772 clade_238 Y OP Bacillus sp. JC6 241 JF824800 clade_238 Y OPBacillus sp. oral taxon F79 248 HM099654 clade_238 Y OP Bacillus sp.SRC_DSF1 243 GU797283 clade_238 Y OP Bacillus sp. SRC_DSF10 242 GU797292clade_238 Y OP Bacillus sp. SRC_DSF2 244 GU797284 clade_238 Y OPBacillus sp. SRC_DSF6 245 GU797288 clade_238 Y OP Bacillus sp. tc09 249HQ844242 clade_238 Y OP Bacillus sp. zh168 250 FJ851424 clade_238 Y OPBacillus sphaericus 251 DQ286318 clade_238 Y OP Bacillussporothermodurans 252 NR_026010 clade_238 Y OP Bacillus subtilis 253EU627588 clade_238 Y OP Bacillus thermoamylovorans 254 NR_029151clade_238 Y OP Bacillus thuringiensis 255 NC_008600 clade_238 Y OPBacillus weihenstephanensis 256 NR_074926 clade_238 Y OP Geobacilluskaustophilus 933 NR_074989 clade_238 Y N Geobacillus stearothermophilus936 NR_040794 clade_238 Y N Geobacillus thermodenitrificans 938NR_074976 clade_238 Y N Geobacillus thermoglucosidasius 939 NR_043022clade_238 Y N Lysinibacillus sphaericus 1193 NR_074883 clade_238 Y NClostridiales sp. SS3_4 543 AY305316 clade_246 Y N Clostridiumbeijerinckii 557 NR_074434 clade_252 Y N Clostridium botulinum 560NC_010723 clade_252 Y Category-A Clostridium butyricum 561 ABDT01000017clade_252 Y N Clostridium chauvoei 568 EU106372 clade_252 Y NClostridium favososporum 582 X76749 clade_252 Y N Clostridiumhistolyticum 592 HF558362 clade_252 Y N Clostridium isatidis 597NR_026347 clade_252 Y N Clostridium limosum 602 FR870444 clade_252 Y NClostridium sartagoforme 622 NR_026490 clade_252 Y N Clostridiumsepticum 624 NR_026020 clade_252 Y N Clostridium sp. 7_2_43FAA 626ACDK01000101 clade_252 Y N Clostridium sporogenes 645 ABKW02000003clade_252 Y N Clostridium tertium 653 Y18174 clade_252 Y N Clostridiumcarnis 564 NR_044716 clade_253 Y N Clostridium celatum 565 X77844clade_253 Y N Clostridium disporicum 579 NR_026491 clade_253 Y NClostridium gasigenes 585 NR_024945 clade_253 Y N Clostridium quinii 616NR_026149 clade_253 Y N Clostridium hylemonae 593 AB023973 clade_260 Y NClostridium scindens 623 AF262238 clade_260 Y N Lachnospiraceaebacterium 5_1_57FAA 1054 ACTR01000020 clade_260 Y N Clostridiumglycyrrhizinilyticum 588 AB233029 clade_262 Y N Clostridium nexile 607X73443 clade_262 Y N Coprococcus comes 674 ABVR01000038 clade_262 Y NLachnospiraceae bacterium 1_1_57FAA 1048 ACTM01000065 clade_262 Y NLachnospiraceae bacterium 1_4_56FAA 1049 ACTN01000028 clade_262 Y NLachnospiraceae bacterium 8_1_57FAA 1057 ACWQ01000079 clade_262 Y NRuminococcus lactaris 1663 ABOU02000049 clade_262 Y N Ruminococcustorques 1670 AAVP02000002 clade_262 Y N Paenibacillus lautus 1397NR_040882 clade_270 Y N Paenibacillus polymyxa 1399 NR_037006 clade_270Y N Paenibacillus sp. HGF5 1402 AEXS01000095 clade_270 Y N Paenibacillussp. HGF7 1403 AFDH01000147 clade_270 Y N Eubacterium sp. oral cloneJI012 868 AY349379 clade_298 Y N Alicyclobacillus contaminans 124NR_041475 clade_301 Y N Alicyclobacillus herbarius 126 NR_024753clade_301 Y N Alicyclobacillus pomorum 127 NR_024801 clade_301 Y NBlautia coccoides 373 AB571656 clade_309 Y N Blautia glucerasea 374AB588023 clade_309 Y N Blautia glucerasei 375 AB439724 clade_309 Y NBlautia hansenii 376 ABYU02000037 clade_309 Y N Blautia luti 378AB691576 clade_309 Y N Blautia producta 379 AB600998 clade_309 Y NBlautia schinkii 380 NR_026312 clade_309 Y N Blautia sp. M25 381HM626178 clade_309 Y N Blautia stercoris 382 HM626177 clade_309 Y NBlautia wexlerae 383 EF036467 clade_309 Y N Bryantella formatexigens 439ACCL02000018 clade_309 Y N Clostridium coccoides 573 EF025906 clade_309Y N Eubacterium cellulosolvens 839 AY178842 clade_309 Y NLachnospiraceae bacterium 6_1_63FAA 1056 ACTV01000014 clade_309 Y NRuminococcus hansenii 1662 M59114 clade_309 Y N Ruminococcus obeum 1664AY169419 clade_309 Y N Ruminococcus sp. 5_1_39BFAA 1666 ACII01000172clade_309 Y N Ruminococcus sp. K_1 1669 AB222208 clade_309 Y NSyntrophococcus sucromutans 1911 NR_036869 clade_309 Y N Bacillusalcalophilus 198 X76436 clade_327 Y N Bacillus clausii 205 FN397477clade_327 Y OP Bacillus gelatini 210 NR_025595 clade_327 Y OP Bacillushalodurans 212 AY144582 clade_327 Y OP Bacillus sp. oral taxon F26 246HM099642 clade_327 Y OP Clostridium innocuum 595 M23732 clade_351 Y NClostridium sp. HGF2 628 AENW01000022 clade_351 Y N Clostridiumperfringens 612 ABDW01000023 clade_353 Y Category-B Sarcina ventriculi1687 NR_026146 clade_353 Y N Clostridium bartlettii 556 ABEZ02000012clade_354 Y N Clostridium bifermentans 558 X73437 clade_354 Y NClostridium ghonii 586 AB542933 clade_354 Y N Clostridium glycolicum 587FJ384385 clade_354 Y N Clostridium mayombei 605 FR733682 clade_354 Y NClostridium sordellii 625 AB448946 clade_354 Y N Clostridium sp. MT4 E635 FJ159523 clade_354 Y N Eubacterium tenue 872 M59118 clade_354 Y NClostridium argentinense 553 NR_029232 clade_355 Y N Clostridium sp.JC122 630 CAEV01000127 clade_355 Y N Clostridium sp. NMBHI_1 636JN093130 clade_355 Y N Clostridium subterminale 650 NR_041795 clade_355Y N Clostridium sulfidigenes 651 NR_044161 clade_355 Y N Doreaformicigenerans 773 AAXA02000006 clade_360 Y N Dorea longicatena 774AJ132842 clade_360 Y N Lachnospiraceae bacterium 2_1_46FAA 1050ADLB01000035 clade_360 Y N Lachnospiraceae bacterium 2_1_58FAA 1051ACTO01000052 clade_360 Y N Lachnospiraceae bacterium 4_1_37FAA 1053ADCR01000030 clade_360 Y N Lachnospiraceae bacterium 9_1_43BFAA 1058ACTX01000023 clade_360 Y N Ruminococcus gnavus 1661 X94967 clade_360 Y NRuminococcus sp. ID8 1668 AY960564 clade_360 Y N Blautiahydrogenotrophica 377 ACBZ01000217 clade_368 Y N Lactonifactorlongoviformis 1147 DQ100449 clade_368 Y N Robinsoniella peoriensis 1633AF445258 clade_368 Y N Eubacterium infirmum 849 U13039 clade_384 Y NEubacterium sp. WAL 14571 864 FJ687606 clade_384 Y N Erysipelotrichaceaebacterium 5_2_54FAA 823 ACZW01000054 clade_385 Y N Eubacterium biforme835 ABYT01000002 clade_385 Y N Eubacterium cylindroides 842 FP929041clade_385 Y N Eubacterium dolichum 844 L34682 clade_385 Y N Eubacteriumsp. 3_1_31 861 ACTL01000045 clade_385 Y N Eubacterium tortuosum 873NR_044648 clade_385 Y N Bulleidia extructa 441 ADFR01000011 clade_388 YN Solobacterium moorei 1739 AECQ01000039 clade_388 Y N Coprococcus catus673 EU266552 clade_393 Y N Lachnospiraceae bacterium oral taxon F15 1064HM099641 clade_393 Y N Clostridium cochlearium 574 NR_044717 clade_395 YN Clostridium malenominatum 604 FR749893 clade_395 Y N Clostridiumtetani 654 NC_004557 clade_395 Y N Acetivibrio ethanolgignens 6 FR749897clade_396 Y N Anaerosporobacter mobilis 161 NR_042953 clade_396 Y NBacteroides pectinophilus 288 ABVQ01000036 clade_396 Y N Clostridiumaminovalericum 551 NR_029245 clade_396 Y N Clostridium phytofermentans613 NR_074652 clade_396 Y N Eubacterium hallii 848 L34621 clade_396 Y NEubacterium xylanophilum 875 L34628 clade_396 Y N Ruminococcus callidus1658 NR_029160 clade_406 Y N Ruminococcus champanellensis 1659 FP929052clade_406 Y N Ruminococcus sp. 18P13 1665 AJ515913 clade_406 Y NRuminococcus sp. 9SE51 1667 FM954974 clade_406 Y N Anaerostipes caccae162 ABAX03000023 clade_408 Y N Anaerostipes sp. 3_2_56FAA 163ACWB01000002 clade_408 Y N Clostridiales bacterium 1_7_47FAA 541ABQR01000074 clade_408 Y N Clostridiales sp. SM4_1 542 FP929060clade_408 Y N Clostridiales sp. SSC_2 544 FP929061 clade_408 Y NClostridium aerotolerans 546 X76163 clade_408 Y N Clostridium aldenense547 NR_043680 clade_408 Y N Clostridium algidixylanolyticum 550NR_028726 clade_408 Y N Clostridium amygdalinum 552 AY353957 clade_408 YN Clostridium asparagiforme 554 ACCJ01000522 clade_408 Y N Clostridiumbolteae 559 ABCC02000039 clade_408 Y N Clostridium celerecrescens 566JQ246092 clade_408 Y N Clostridium citroniae 569 ADLJ01000059 clade_408Y N Clostridium clostridiiformes 571 M59089 clade_408 Y N Clostridiumclostridioforme 572 NR_044715 clade_408 Y N Clostridium hathewayi 590AY552788 clade_408 Y N Clostridium indolis 594 AF028351 clade_408 Y NClostridium lavalense 600 EF564277 clade_408 Y N Clostridiumsaccharolyticum 620 CP002109 clade_408 Y N Clostridium sp. M62_1 633ACFX02000046 clade_408 Y N Clostridium sp. SS2_1 638 ABGC03000041clade_408 Y N Clostridium sphenoides 643 X73449 clade_408 Y NClostridium symbiosum 652 ADLQ01000114 clade_408 Y N Clostridiumxylanolyticum 658 NR_037068 clade_408 Y N Eubacterium hadrum 847FR749933 clade_408 Y N Lachnospiraceae bacterium 3_1_57FAA_CT1 1052ACTP01000124 clade_408 Y N Lachnospiraceae bacterium 5_1_63FAA 1055ACTS01000081 clade_408 Y N Lachnospiraceae bacterium A4 1059 DQ789118clade_408 Y N Lachnospiraceae bacterium DJF VP30 1060 EU728771 clade_408Y N Lachnospiraceae genomosp. C1 1065 AY278618 clade_408 Y N Clostridiumdifficile 578 NC_013315 clade_409 Y OP Eubacterium sp. AS15b 862HQ616364 clade_428 Y N Eubacterium sp. OBRC9 863 HQ616354 clade_428 Y NEubacterium sp. oral clone OH3A 871 AY947497 clade_428 Y N Eubacteriumyurii 876 AEES01000073 clade_428 Y N Clostridium acetobutylicum 545NR_074511 clade_430 Y N Clostridium algidicarnis 549 NR_041746 clade_430Y N Clostridium cadaveris 562 AB542932 clade_430 Y N Clostridiumcarboxidivorans 563 FR733710 clade_430 Y N Clostridium estertheticum 580NR_042153 clade_430 Y N Clostridium fallax 581 NR_044714 clade_430 Y NClostridium felsineum 583 AF270502 clade_430 Y N Clostridiumfrigidicarnis 584 NR_024919 clade_430 Y N Clostridium kluyveri 598NR_074165 clade_430 Y N Clostridium magnum 603 X77835 clade_430 Y NClostridium putrefaciens 615 NR_024995 clade_430 Y N Clostridium sp.HPB_46 629 AY862516 clade_430 Y N Clostridium tyrobutyricum 656NR_044718 clade_430 Y N Sutterella parvirubra 1899 AB300989 clade_432 YN Acetanaerobacterium elongatum 4 NR_042930 clade_439 Y N Clostridiumcellulosi 567 NR_044624 clade_439 Y N Ethanoligenens harbinense 832AY675965 clade_439 Y N Eubacterium rectale 856 FP929042 clade_444 Y NEubacterium sp. oral clone GI038 865 AY349374 clade_444 Y NLachnobacterium bovis 1045 GU324407 clade_444 Y N Roseburia cecicola1634 GU233441 clade_444 Y N Roseburia faecalis 1635 AY804149 clade_444 YN Roseburia faecis 1636 AY305310 clade_444 Y N Roseburia hominis 1637AJ270482 clade_444 Y N Roseburia intestinalis 1638 FP929050 clade_444 YN Roseburia inulinivorans 1639 AJ270473 clade_444 Y N Brevibacillusbrevis 410 NR_041524 clade_448 Y N Brevibacillus laterosporus 414NR_037005 clade_448 Y N Bacillus coagulans 206 DQ297928 clade_451 Y OPSporolactobacillus inulinus 1752 NR_040962 clade_451 Y N Kocuriapalustris 1041 EU333884 clade_453 Y N Nocardia farcinica 1353 NC_006361clade_455 Y N Bacillus sp. oral taxon F28 247 HM099650 clade_456 Y OPCatenibacterium mitsuokai 495 AB030224 clade_469 Y N Clostridium sp.TM_40 640 AB249652 clade_469 Y N Coprobacillus cateniformis 670 AB030218clade_469 Y N Coprobacillus sp. 29_1 671 ADKX01000057 clade_469 Y NClostridium rectum 618 NR_029271 clade_470 Y N Eubacterium nodatum 854U13041 clade_476 Y N Eubacterium saphenum 859 NR_026031 clade_476 Y NEubacterium sp. oral clone JH012 867 AY349373 clade_476 Y N Eubacteriumsp. oral clone JS001 870 AY349378 clade_476 Y N Faecalibacteriumprausnitzii 880 ACOP02000011 clade_478 Y N Gemmiger formicilis 932GU562446 clade_478 Y N Subdoligranulum variabile 1896 AJ518869 clade_478Y N Clostridiaceae bacterium JC13 532 JF824807 clade_479 Y N Clostridiumsp. MLG055 634 AF304435 clade_479 Y N Erysipelotrichaceae bacterium3_1_53 822 ACTJ01000113 clade_479 Y N Clostridium cocleatum 575NR_026495 clade_481 Y N Clostridium ramosum 617 M23731 clade_481 Y NClostridium saccharogumia 619 DQ100445 clade_481 Y N Clostridiumspiroforme 644 X73441 clade_481 Y N Coprobacillus sp. D7 672ACDT01000199 clade_481 Y N Clostridiales bacterium SY8519 535 AB477431clade_482 Y N Clostridium sp. SY8519 639 AP012212 clade_482 Y NEubacterium ramulus 855 AJ011522 clade_482 Y N Erysipelothrix inopinata819 NR_025594 clade_485 Y N Erysipelothrix rhusiopathiae 820ACLK01000021 clade_485 Y N Erysipelothrix tonsillarum 821 NR_040871clade_485 Y N Holdemania filiformis 1004 Y11466 clade_485 Y N Mollicutesbacterium pACH93 1258 AY297808 clade_485 Y N Coxiella burnetii 736CP000890 clade_486 Y Category-B Clostridium hiranonis 591 AB023970clade_487 Y N Clostridium irregulare 596 NR_029249 clade_487 Y NClostridium orbiscindens 609 Y18187 clade_494 Y N Clostridium sp. NML04A032 637 EU815224 clade_494 Y N Flavonifractor plautii 886 AY724678clade_494 Y N Pseudoflavonifractor capillosus 1591 AY136666 clade_494 YN Ruminococcaceae bacterium D16 1655 ADDX01000083 clade_494 Y NAcetivibrio cellulolyticus 5 NR_025917 clade_495 Y N Clostridiumaldrichii 548 NR_026099 clade_495 Y N Clostridium clariflavum 570NR_041235 clade_495 Y N Clostridium stercorarium 647 NR_025100 clade_495Y N Clostridium straminisolvens 649 NR_024829 clade_495 Y N Clostridiumthermocellum 655 NR_074629 clade_495 Y N Fusobacterium nucleatum 901ADVK01000034 clade_497 Y N Eubacterium barkeri 834 NR_044661 clade_512 YN Eubacterium callanderi 838 NR_026330 clade_512 Y N Eubacterium limosum850 CP002273 clade_512 Y N Anaerotruncus colihominis 164 ABGD02000021clade_516 Y N Clostridium methylpentosum 606 ACEC01000059 clade_516 Y NClostridium sp. YIT 12070 642 AB491208 clade_516 Y NHydrogenoanaerobacterium saccharovorans 1005 NR_044425 clade_516 Y NRuminococcus albus 1656 AY445600 clade_516 Y N Ruminococcus flavefaciens1660 NR_025931 clade_516 Y N Clostridium haemolyticum 589 NR_024749clade_517 Y N Clostridium novyi 608 NR_074343 clade_517 Y N Clostridiumsp. LMG 16094 632 X95274 clade_517 Y N Eubacterium ventriosum 874 L34421clade_519 Y N Bacteroides galacturonicus 280 DQ497994 clade_522 Y NEubacterium eligens 845 CP001104 clade_522 Y N Lachnospira multipara1046 FR733699 clade_522 Y N Lachnospira pectinoschiza 1047 L14675clade_522 Y N Lactobacillus rogosae 1114 GU269544 clade_522 Y N Bacillushorti 214 NR_036860 clade_527 Y OP Bacillus sp. 9_3AIA 232 FN397519clade_527 Y OP Eubacterium brachy 836 U13038 clade_533 Y N Filifactoralocis 881 CP002390 clade_533 Y N Filifactor villosus 882 NR_041928clade_533 Y N Clostridium leptum 601 AJ305238 clade_537 Y N Clostridiumsp. YIT 12069 641 AB491207 clade_537 Y N Clostridium sporosphaeroides646 NR_044835 clade_537 Y N Eubacterium coprostanoligenes 841 HM037995clade_537 Y N Ruminococcus bromii 1657 EU266549 clade_537 Y NEubacterium siraeum 860 ABCA03000054 clade_538 Y N Clostridium viride657 NR_026204 clade_540 Y N Oscillibacter sp. G2 1386 HM626173 clade_540Y N Oscillibacter valericigenes 1387 NR_074793 clade_540 Y NOscillospira guilliermondii 1388 AB040495 clade_540 Y N Butyrivibriocrossotus 455 ABWN01000012 clade_543 Y N Clostridium sp. L2_50 631AAYW02000018 clade_543 Y N Coprococcus eutactus 675 EF031543 clade_543 YN Coprococcus sp. ART55_1 676 AY350746 clade_543 Y N Eubacteriumruminantium 857 NR_024661 clade_543 Y N Collinsella aerofaciens 659AAVN02000007 clade_553 Y N Alkaliphilus metalliredigenes 137 AY137848clade_554 Y N Alkaliphilus oremlandii 138 NR_043674 clade_554 Y NClostridium sticklandii 648 L04167 clade_554 Y N Turicibacter sanguinis1965 AF349724 clade_555 Y N Fulvimonas sp. NML 060897 892 EF589680clade_557 Y N Desulfitobacterium frappieri 753 AJ276701 clade_560 Y NDesulfitobacterium hafniense 754 NR_074996 clade_560 Y NDesulfotomaculum nigrificans 756 NR_044832 clade_560 Y N Lutisporathermophila 1191 NR_041236 clade_564 Y N Brachyspira pilosicoli 405NR_075069 clade_565 Y N Eggerthella lenta 778 AF292375 clade_566 Y NStreptomyces albus 1888 AJ697941 clade_566 Y N Chlamydiales bacteriumNS11 505 JN606074 clade_567 Y N Anaerofustis stercorihominis 159ABIL02000005 clade_570 Y N Butyricicoccus pullicaecorum 453 HH793440clade_572 Y N Eubacterium desmolans 843 NR_044644 clade_572 Y NPapillibacter cinnamivorans 1415 NR_025025 clade_572 Y N Sporobactertermitidis 1751 NR_044972 clade_572 Y N Deferribacteres sp. oral cloneJV006 744 AY349371 clade_575 Y N Clostridium colinum 577 NR_026151clade_576 Y N Clostridium lactatifermentans 599 NR_025651 clade_576 Y NClostridium piliforme 614 D14639 clade_576 Y N Saccharomonospora viridis1671 X54286 clade_579 Y N Thermobifida fusca 1921 NC_007333 clade_579 YN Leptospira licerasiae 1164 EF612284 clade_585 Y OP Moorellathermoacetica 1259 NR_075001 clade_590 Y N Thermoanaerobacterpseudethanolicus 1920 CP000924 clade_590 Y N Flexistipes sinusarabici888 NR_074881 clade_591 Y N Gloeobacter violaceus 942 NR_074282clade_596 Y N Eubacterium sp. oral clone JN088 869 AY349377 clade_90 Y NClostridium oroticum 610 FR749922 clade_96 Y N Clostridium sp. D5 627ADBG01000142 clade_96 Y N Eubacterium contortum 840 FR749946 clade_96 YN Eubacterium fissicatena 846 FR749935 clade_96 Y N Corynebacteriumcoyleae 692 X96497 clade_100 N N Corynebacterium mucifaciens 711NR_026396 clade_100 N N Corynebacterium ureicelerivorans 733 AM397636clade_100 N N Corynebacterium appendicis 684 NR_028951 clade_102 N NCorynebacterium genitalium 698 ACLJ01000031 clade_102 N NCorynebacterium glaucum 699 NR_028971 clade_102 N N Corynebacteriumimitans 703 AF537597 clade_102 N N Corynebacterium riegelii 719 EU848548clade_102 N N Corynebacterium sp. L_2012475 723 HE575405 clade_102 N NCorynebacterium sp. NML 93_0481 724 GU238409 clade_102 N NCorynebacterium sundsvallense 728 Y09655 clade_102 N N Corynebacteriumtuscaniae 730 AY677186 clade_102 N N Prevotella maculosa 1504AGEK01000035 clade_104 N N Prevotella oris 1513 ADDV01000091 clade_104 NN Prevotella salivae 1517 AB108826 clade_104 N N Prevotella sp. ICM551521 HQ616399 clade_104 N N Prevotella sp. oral clone AA020 1528AY005057 clade_104 N N Prevotella sp. oral clone GI032 1538 AY349396clade_104 N N Prevotella sp. oral taxon G70 1558 GU432179 clade_104 N NPrevotella corporis 1491 L16465 clade_105 N N Bacteroides sp. 4_1_36 312ACTC01000133 clade_110 N N Bacteroides sp. AR20 315 AF139524 clade_110 NN Bacteroides sp. D20 319 ACPT01000052 clade_110 N N Bacteroides sp. F_4322 AB470322 clade_110 N N Bacteroides uniformis 329 AB050110 clade_110N N Prevotella nanceiensis 1510 JN867228 clade_127 N N Prevotella sp.oral taxon 299 1548 ACWZ01000026 clade_127 N N Prevotella bergensis 1485ACKS01000100 clade_128 N N Prevotella buccalis 1489 JN867261 clade_129 NN Prevotella timonensis 1564 ADEF01000012 clade_129 N N Prevotellaoralis 1512 AEPE01000021 clade_130 N N Prevotella sp. SEQ072 1525JN867238 clade_130 N N Leuconostoc carnosum 1177 NR_040811 clade_135 N NLeuconostoc gasicomitatum 1179 FN822744 clade_135 N N Leuconostoc inhae1180 NR_025204 clade_135 N N Leuconostoc kimchii 1181 NR_075014clade_135 N N Edwardsiella tarda 777 CP002154 clade_139 N N Photorhabdusasymbiotica 1466 Z76752 clade_139 N N Psychrobacter arcticus 1607CP000082 clade_141 N N Psychrobacter cibarius 1608 HQ698586 clade_141 NN Psychrobacter cryohalolentis 1609 CP000323 clade_141 N N Psychrobacterfaecalis 1610 HQ698566 clade_141 N N Psychrobacter nivimaris 1611HQ698587 clade_141 N N Psychrobacter pulmonis 1612 HQ698582 clade_141 NN Pseudomonas aeruginosa 1592 AABQ07000001 clade_154 N N Pseudomonas sp.2_1_26 1600 ACWU01000257 clade_154 N N Corynebacterium confusum 691Y15886 clade_158 N N Corynebacterium propinquum 712 NR_037038 clade_158N N Corynebacterium pseudodiphtheriticum 713 X84258 clade_158 N NBartonella bacilliformis 338 NC_008783 clade_159 N N Bartonella grahamii339 CP001562 clade_159 N N Bartonella henselae 340 NC_005956 clade_159 NN Bartonella quintana 341 BX897700 clade_159 N N Bartonella tamiae 342EF672728 clade_159 N N Bartonella washoensis 343 FJ719017 clade_159 N NBrucella abortus 430 ACBJ01000075 clade_159 N Category-B Brucella canis431 NR_044652 clade_159 N Category-B Brucella ceti 432 ACJD01000006clade_159 N Category-B Brucella melitensis 433 AE009462 clade_159 NCategory-B Brucella microti 434 NR_042549 clade_159 N Category-BBrucella ovis 435 NC_009504 clade_159 N Category-B Brucella sp. 83_13436 ACBQ01000040 clade_159 N Category-B Brucella sp. BO1 437 EU053207clade_159 N Category-B Brucella suis 438 ACBK01000034 clade_159 NCategory-B Ochrobactrum anthropi 1360 NC_009667 clade_159 N NOchrobactrum intermedium 1361 ACQA01000001 clade_159 N N Ochrobactrumpseudintermedium 1362 DQ365921 clade_159 N N Prevotella genomosp. C21496 AY278625 clade_164 N N Prevotella multisaccharivorax 1509AFJE01000016 clade_164 N N Prevotella sp. oral clone IDR_CEC_0055 1543AY550997 clade_164 N N Prevotella sp. oral taxon 292 1547 GQ422735clade_164 N N Prevotella sp. oral taxon 300 1549 GU409549 clade_164 N NPrevotella marshii 1505 AEEI01000070 clade_166 N N Prevotella sp. oralclone IK053 1544 AY349401 clade_166 N N Prevotella sp. oral taxon 7811554 GQ422744 clade_166 N N Prevotella stercorea 1562 AB244774 clade_166N N Prevotella brevis 1487 NR_041954 clade_167 N N Prevotella ruminicola1516 CP002006 clade_167 N N Prevotella sp. sp24 1560 AB003384 clade_167N N Prevotella sp. sp34 1561 AB003385 clade_167 N N Prevotella albensis1483 NR_025300 clade_168 N N Prevotella copri 1490 ACBX02000014clade_168 N N Prevotella oulorum 1514 L16472 clade_168 N N Prevotellasp. BI_42 1518 AJ581354 clade_168 N N Prevotella sp. oral clone P4PB_83P2 1546 AY207050 clade_168 N N Prevotella sp. oral taxon G60 1557GU432133 clade_168 N N Prevotella amnii 1484 AB547670 clade_169 N NBacteroides caccae 268 EU136686 clade_170 N N Bacteroides finegoldii 277AB222699 clade_170 N N Bacteroides intestinalis 283 ABJL02000006clade_171 N N Bacteroides sp. XB44A 326 AM230649 clade_171 N NBifidobacteriaceae genomosp. C1 345 AY278612 clade_172 N NBifidobacterium adolescentis 346 AAXD02000018 clade_172 N NBifidobacterium angulatum 347 ABYS02000004 clade_172 N N Bifidobacteriumanimalis 348 CP001606 clade_172 N N Bifidobacterium breve 350 CP002743clade_172 N N Bifidobacterium catenulatum 351 ABXY01000019 clade_172 N NBifidobacterium dentium 352 CP001750 clade_172 N OP Bifidobacteriumgallicum 353 ABXB03000004 clade_172 N N Bifidobacterium infantis 354AY151398 clade_172 N N Bifidobacterium kashiwanohense 355 AB491757clade_172 N N Bifidobacterium longum 356 ABQQ01000041 clade_172 N NBifidobacterium pseudocatenulatum 357 ABXX02000002 clade_172 N NBifidobacterium pseudolongum 358 NR_043442 clade_172 N N Bifidobacteriumscardovii 359 AJ307005 clade_172 N N Bifidobacterium sp. HM2 360AB425276 clade_172 N N Bifidobacterium sp. HMLN12 361 JF519685 clade_172N N Bifidobacterium sp. M45 362 HM626176 clade_172 N N Bifidobacteriumsp. MSX5B 363 HQ616382 clade_172 N N Bifidobacterium sp. TM_7 364AB218972 clade_172 N N Bifidobacterium thermophilum 365 DQ340557clade_172 N N Leuconostoc citreum 1178 AM157444 clade_175 N NLeuconostoc lactis 1182 NR_040823 clade_175 N N Alicyclobacillusacidoterrestris 123 NR_040844 clade_179 N N Alicyclobacilluscycloheptanicus 125 NR_024754 clade_179 N N Acinetobacter baumannii 27ACYQ01000014 clade_181 N N Acinetobacter calcoaceticus 28 AM157426clade_181 N N Acinetobacter genomosp. C1 29 AY278636 clade_181 N NAcinetobacter haemolyticus 30 ADMT01000017 clade_181 N N Acinetobacterjohnsonii 31 ACPL01000162 clade_181 N N Acinetobacter junii 32ACPM01000135 clade_181 N N Acinetobacter lwoffii 33 ACPN01000204clade_181 N N Acinetobacter parvus 34 AIEB01000124 clade_181 N NAcinetobacter schindleri 36 NR_025412 clade_181 N N Acinetobacter sp.56A1 37 GQ178049 clade_181 N N Acinetobacter sp. CIP 101934 38 JQ638573clade_181 N N Acinetobacter sp. CIP 102143 39 JQ638578 clade_181 N NAcinetobacter sp. M16_22 41 HM366447 clade_181 N N Acinetobacter sp.RUH2624 42 ACQF01000094 clade_181 N N Acinetobacter sp. SH024 43ADCH01000068 clade_181 N N Lactobacillus jensenii 1092 ACQD01000066clade_182 N N Alcaligenes faecalis 119 AB680368 clade_183 N NAlcaligenes sp. CO14 120 DQ643040 clade_183 N N Alcaligenes sp. S3 121HQ262549 clade_183 N N Oligella ureolytica 1366 NR_041998 clade_183 N NOligella urethralis 1367 NR_041753 clade_183 N N Eikenella corrodens 784ACEA01000028 clade_185 N N Kingella denitrificans 1019 AEWV01000047clade_185 N N Kingella genomosp. P1 oral cone MB2_C20 1020 DQ003616clade_185 N N Kingella kingae 1021 AFHS01000073 clade_185 N N Kingellaoralis 1022 ACJW02000005 clade_185 N N Kingella sp. oral clone ID0591023 AY349381 clade_185 N N Neisseria elongata 1330 ADBF01000003clade_185 N N Neisseria genomosp. P2 oral clone MB5_P15 1332 DQ003630clade_185 N N Neisseria sp. oral clone JC012 1345 AY349388 clade_185 N NNeisseria sp. SMC_A9199 1342 FJ763637 clade_185 N N Simonsiella muelleri1731 ADCY01000105 clade_185 N N Corynebacterium glucuronolyticum 700ABYP01000081 clade_193 N N Corynebacterium pyruviciproducens 716FJ185225 clade_193 N N Rothia aeria 1649 DQ673320 clade_194 N N Rothiadentocariosa 1650 ADDW01000024 clade_194 N N Rothia sp. oral taxon 1881653 GU470892 clade_194 N N Corynebacterium accolens 681 ACGD01000048clade_195 N N Corynebacterium macginleyi 707 AB359393 clade_195 N NCorynebacterium pseudogenitalium 714 ABYQ01000237 clade_195 N NCorynebacterium tuberculostearicum 729 ACVP01000009 clade_195 N NLactobacillus casei 1074 CP000423 clade_198 N N Lactobacillus paracasei1106 ABQV01000067 clade_198 N N Lactobacillus zeae 1143 NR_037122clade_198 N N Prevotella dentalis 1492 AB547678 clade_205 N N Prevotellasp. oral clone ASCG10 1529 AY923148 clade_206 N N Prevotella sp. oralclone HF050 1541 AY349399 clade_206 N N Prevotella sp. oral clone ID0191542 AY349400 clade_206 N N Prevotella sp. oral clone IK062 1545AY349402 clade_206 N N Prevotella genomosp. P9 oral clone MB7_G16 1499DQ003633 clade_207 N N Prevotella sp. oral clone AU069 1531 AY005062clade_207 N N Prevotella sp. oral clone CY006 1532 AY005063 clade_207 NN Prevotella sp. oral clone FL019 1534 AY349392 clade_207 N NActinomyces genomosp. C1 56 AY278610 clade_212 N N Actinomyces genomosp.C2 57 AY278611 clade_212 N N Actinomyces genomosp. P1 oral clone MB6_C0358 DQ003632 clade_212 N N Actinomyces georgiae 59 GU561319 clade_212 N NActinomyces israelii 60 AF479270 clade_212 N N Actinomyces massiliensis61 AB545934 clade_212 N N Actinomyces meyeri 62 GU561321 clade_212 N NActinomyces odontolyticus 66 ACYT01000123 clade_212 N N Actinomycesorihominis 68 AJ575186 clade_212 N N Actinomyces sp. CCUG 37290 71AJ234058 clade_212 N N Actinomyces sp. ICM34 75 HQ616391 clade_212 N NActinomyces sp. ICM41 76 HQ616392 clade_212 N N Actinomyces sp. ICM47 77HQ616395 clade_212 N N Actinomyces sp. ICM54 78 HQ616398 clade_212 N NActinomyces sp. oral clone IP081 87 AY349366 clade_212 N N Actinomycessp. oral taxon 178 91 AEUH01000060 clade_212 N N Actinomyces sp. oraltaxon 180 92 AEPP01000041 clade_212 N N Actinomyces sp. TeJ5 80 GU561315clade_212 N N Haematobacter sp. BC14248 968 GU396991 clade_213 N NParacoccus denitrificans 1424 CP000490 clade_213 N N Paracoccus marcusii1425 NR_044922 clade_213 N N Grimontia hollisae 967 ADAQ01000013clade_216 N N Shewanella putrefaciens 1723 CP002457 clade_216 N N Afipiagenomosp. 4 111 EU117385 clade_217 N N Rhodopseudomonas palustris 1626CP000301 clade_217 N N Methylobacterium extorquens 1223 NC_010172clade_218 N N Methylobacterium podarium 1224 AY468363 clade_218 N NMethylobacterium radiotolerans 1225 GU294320 clade_218 N NMethylobacterium sp. 1sub 1226 AY468371 clade_218 N N Methylobacteriumsp. MM4 1227 AY468370 clade_218 N N Achromobacter denitrificans 18NR_042021 clade_224 N N Achromobacter piechaudii 19 ADMS01000149clade_224 N N Achromobacter xylosoxidans 20 ACRC01000072 clade_224 N NBordetella bronchiseptica 384 NR_025949 clade_224 N OP Bordetellaholmesii 385 AB683187 clade_224 N OP Bordetella parapertussis 386NR_025950 clade_224 N OP Bordetella pertussis 387 BX640418 clade_224 NOP Microbacterium chocolatum 1230 NR_037045 clade_225 N N Microbacteriumflavescens 1231 EU714363 clade_225 N N Microbacterium lacticum 1233EU714351 clade_225 N N Microbacterium oleivorans 1234 EU714381 clade_225N N Microbacterium oxydans 1235 EU714348 clade_225 N N Microbacteriumparaoxydans 1236 AJ491806 clade_225 N N Microbacterium phyllosphaerae1237 EU714359 clade_225 N N Microbacterium schleiferi 1238 NR_044936clade_225 N N Microbacterium sp. 768 1239 EU714378 clade_225 N NMicrobacterium sp. oral strain C24KA 1240 AF287752 clade_225 N NMicrobacterium testaceum 1241 EU714365 clade_225 N N Corynebacteriumatypicum 686 NR_025540 clade_229 N N Corynebacterium mastitidis 708AB359395 clade_229 N N Corynebacterium sp. NML 97_0186 725 GU238411clade_229 N N Mycobacterium elephantis 1275 AF385898 clade_237 N OPMycobacterium paraterrae 1288 EU919229 clade_237 N OP Mycobacteriumphlei 1289 GU142920 clade_237 N OP Mycobacterium sp. 1776 1293 EU703152clade_237 N N Mycobacterium sp. 1781 1294 EU703147 clade_237 N NMycobacterium sp. AQ1GA4 1297 HM210417 clade_237 N N Mycobacterium sp.GN_10546 1299 FJ497243 clade_237 N N Mycobacterium sp. GN_10827 1300FJ497247 clade_237 N N Mycobacterium sp. GN_11124 1301 FJ652846clade_237 N N Mycobacterium sp. GN_9188 1302 FJ497240 clade_237 N NMycobacterium sp. GR_2007_210 1303 FJ555538 clade_237 N N Anoxybacilluscontaminans 172 NR_029006 clade_238 N N Bacillus aeolius 195 NR_025557clade_238 N N Brevibacterium frigoritolerans 422 NR_042639 clade_238 N NGeobacillus sp. E263 934 DQ647387 clade_238 N N Geobacillus sp. WCH70935 CP001638 clade_238 N N Geobacillus thermocatenulatus 937 NR_043020clade_238 N N Geobacillus thermoleovorans 940 NR_074931 clade_238 N NLysinibacillus fusiformis 1192 FN397522 clade_238 N N Planomicrobiumkoreense 1468 NR_025011 clade_238 N N Sporosarcina newyorkensis 1754AFPZ01000142 clade_238 N N Sporosarcina sp. 2681 1755 GU994081 clade_238N N Ureibacillus composti 1968 NR_043746 clade_238 N N Ureibacillussuwonensis 1969 NR_043232 clade_238 N N Ureibacillus terrenus 1970NR_025394 clade_238 N N Ureibacillus thermophilus 1971 NR_043747clade_238 N N Ureibacillus thermosphaericus 1972 NR_040961 clade_238 N NPrevotella micans 1507 AGWK01000061 clade_239 N N Prevotella sp. oralclone DA058 1533 AY005065 clade_239 N N Prevotella sp. SEQ053 1523JN867222 clade_239 N N Treponema socranskii 1937 NR_024868 clade_240 NOP Treponema sp. 6:H:D15A_4 1938 AY005083 clade_240 N N Treponema sp.oral taxon 265 1953 GU408850 clade_240 N N Treponema sp. oral taxon G851958 GU432215 clade_240 N N Porphyromonas endodontalis 1472 ACNN01000021clade_241 N N Porphyromonas sp. oral clone BB134 1478 AY005068 clade_241N N Porphyromonas sp. oral clone F016 1479 AY005069 clade_241 N NPorphyromonas sp. oral clone P2PB_52 P1 1480 AY207054 clade_241 N NPorphyromonas sp. oral clone P4GB_100 P2 1481 AY207057 clade_241 N NAcidovorax sp. 98_63833 26 AY258065 clade_245 N N Comamonadaceaebacterium NML000135 663 JN585335 clade_245 N N Comamonadaceae bacteriumNML790751 664 JN585331 clade_245 N N Comamonadaceae bacterium NML910035665 JN585332 clade_245 N N Comamonadaceae bacterium NML910036 666JN585333 clade_245 N N Comamonas sp. NSP5 668 AB076850 clade_245 N NDelftia acidovorans 748 CP000884 clade_245 N N Xenophilus aerolatus 2018JN585329 clade_245 N N Oribacterium sp. oral taxon 078 1380 ACIQ02000009clade_246 N N Oribacterium sp. oral taxon 102 1381 GQ422713 clade_246 NN Weissella cibaria 2007 NR_036924 clade_247 N N Weissella confusa 2008NR_040816 clade_247 N N Weissella hellenica 2009 AB680902 clade_247 N NWeissella kandleri 2010 NR_044659 clade_247 N N Weissella koreensis 2011NR_075058 clade_247 N N Weissella paramesenteroides 2012 ACKU01000017clade_247 N N Weissella sp. KLDS 7.0701 2013 EU600924 clade_247 N NMobiluncus curtisii 1251 AEPZ01000013 clade_249 N N Enhydrobacteraerosaccus 785 ACYI01000081 clade_256 N N Moraxella osloensis 1262JN175341 clade_256 N N Moraxella sp. GM2 1264 JF837191 clade_256 N NBrevibacterium casei 420 JF951998 clade_257 N N Brevibacteriumepidermidis 421 NR_029262 clade_257 N N Brevibacterium sanguinis 426NR_028016 clade_257 N N Brevibacterium sp. H15 427 AB177640 clade_257 NN Acinetobacter radioresistens 35 ACVR01000010 clade_261 N NLactobacillus alimentarius 1068 NR_044701 clade_263 N N Lactobacillusfarciminis 1082 NR_044707 clade_263 N N Lactobacillus kimchii 1097NR_025045 clade_263 N N Lactobacillus nodensis 1101 NR_041629 clade_263N N Lactobacillus tucceti 1138 NR_042194 clade_263 N N Pseudomonasmendocina 1595 AAUL01000021 clade_265 N N Pseudomonas pseudoalcaligenes1598 NR_037000 clade_265 N N Pseudomonas sp. NP522b 1602 EU723211clade_265 N N Pseudomonas stutzeri 1603 AM905854 clade_265 N NPaenibacillus barcinonensis 1390 NR_042272 clade_270 N N Paenibacillusbarengoltzii 1391 NR_042756 clade_270 N N Paenibacillus chibensis 1392NR_040885 clade_270 N N Paenibacillus cookii 1393 NR_025372 clade_270 NN Paenibacillus durus 1394 NR_037017 clade_270 N N Paenibacillusglucanolyticus 1395 D78470 clade_270 N N Paenibacillus lactis 1396NR_025739 clade_270 N N Paenibacillus pabuli 1398 NR_040853 clade_270 NN Paenibacillus popilliae 1400 NR_040888 clade_270 N N Paenibacillus sp.CIP 101062 1401 HM212646 clade_270 N N Paenibacillus sp. JC66 1404JF824808 clade_270 N N Paenibacillus sp. R_27413 1405 HE586333 clade_270N N Paenibacillus sp. R_27422 1406 HE586338 clade_270 N N Paenibacillustimonensis 1408 NR_042844 clade_270 N N Rothia mucilaginosa 1651ACVO01000020 clade_271 N N Rothia nasimurium 1652 NR_025310 clade_271 NN Prevotella sp. oral taxon 302 1550 ACZK01000043 clade_280 N NPrevotella sp. oral taxon F68 1556 HM099652 clade_280 N N Prevotellatannerae 1563 ACIJ02000018 clade_280 N N Prevotellaceae bacterium P4P_62P1 1566 AY207061 clade_280 N N Porphyromonas asaccharolytica 1471AENO01000048 clade_281 N N Porphyromonas gingivalis 1473 AE015924clade_281 N N Porphyromonas macacae 1475 NR_025908 clade_281 N NPorphyromonas sp. UQD 301 1477 EU012301 clade_281 N N Porphyromonasuenonis 1482 ACLR01000152 clade_281 N N Leptotrichia buccalis 1165CP001685 clade_282 N N Leptotrichia hofstadii 1168 ACVB02000032clade_282 N N Leptotrichia sp. oral clone HE012 1173 AY349386 clade_282N N Leptotrichia sp. oral taxon 223 1176 GU408547 clade_282 N NBacteroides fluxus 278 AFBN01000029 clade_285 N N Bacteroides helcogenes281 CP002352 clade_285 N N Parabacteroides johnsonii 1419 ABYH01000014clade_286 N N Parabacteroides merdae 1420 EU136685 clade_286 N NTreponema denticola 1926 ADEC01000002 clade_288 N OP Treponema genomosp.P5 oral clone MB3_P23 1929 DQ003624 clade_288 N N Treponema putidum 1935AJ543428 clade_288 N OP Treponema sp. oral clone P2PB_53 P3 1942AY207055 clade_288 N N Treponema sp. oral taxon 247 1949 GU408748clade_288 N N Treponema sp. oral taxon 250 1950 GU408776 clade_288 N NTreponema sp. oral taxon 251 1951 GU408781 clade_288 N N Anaerococcushydrogenalis 144 ABXA01000039 clade_289 N N Anaerococcus sp. 8404299 148HM587318 clade_289 N N Anaerococcus sp. gpac215 156 AM176540 clade_289 NN Anaerococcus vaginalis 158 ACXU01000016 clade_289 N NPropionibacterium acidipropionici 1569 NC_019395 clade_290 N NPropionibacterium avidum 1571 AJ003055 clade_290 N N Propionibacteriumgranulosum 1573 FJ785716 clade_290 N N Propionibacterium jensenii 1574NR_042269 clade_290 N N Propionibacterium propionicum 1575 NR_025277clade_290 N N Propionibacterium sp. H456 1577 AB177643 clade_290 N NPropionibacterium thoenii 1581 NR_042270 clade_290 N N Bifidobacteriumbifidum 349 ABQP01000027 clade_293 N N Leuconostoc mesenteroides 1183ACKV01000113 clade_295 N N Leuconostoc pseudomesenteroides 1184NR_040814 clade_295 N N Johnsonella ignava 1016 X87152 clade_298 N NPropionibacterium acnes 1570 ADJM01000010 clade_299 N NPropionibacterium sp. 434_HC2 1576 AFIL01000035 clade_299 N NPropionibacterium sp. LG 1578 AY354921 clade_299 N N Propionibacteriumsp. S555a 1579 AB264622 clade_299 N N Alicyclobacillus sp. CCUG 53762128 HE613268 clade_301 N N Actinomyces cardiffensis 53 GU470888clade_303 N N Actinomyces funkei 55 HQ906497 clade_303 N N Actinomycessp. HKU31 74 HQ335393 clade_303 N N Actinomyces sp. oral taxon C55 94HM099646 clade_303 N N Kerstersia gyiorum 1018 NR_025669 clade_307 N NPigmentiphaga daeguensis 1467 JN585327 clade_307 N N Aeromonasallosaccharophila 104 S39232 clade_308 N N Aeromonas enteropelogenes 105X71121 clade_308 N N Aeromonas hydrophila 106 NC_008570 clade_308 N NAeromonas jandaei 107 X60413 clade_308 N N Aeromonas salmonicida 108NC_009348 clade_308 N N Aeromonas trota 109 X60415 clade_308 N NAeromonas veronii 110 NR_044845 clade_308 N N Marvinbryantiaformatexigens 1196 AJ505973 clade_309 N N Rhodobacter sp. oral taxon C301620 HM099648 clade_310 N N Rhodobacter sphaeroides 1621 CP000144clade_310 N N Lactobacillus antri 1071 ACLL01000037 clade_313 N NLactobacillus coleohominis 1076 ACOH01000030 clade_313 N N Lactobacillusfermentum 1083 CP002033 clade_313 N N Lactobacillus gastricus 1085AICN01000060 clade_313 N N Lactobacillus mucosae 1099 FR693800 clade_313N N Lactobacillus oris 1103 AEKL01000077 clade_313 N N Lactobacilluspontis 1111 HM218420 clade_313 N N Lactobacillus reuteri 1112ACGW02000012 clade_313 N N Lactobacillus sp. KLDS 1.0707 1127 EU600911clade_313 N N Lactobacillus sp. KLDS 1.0709 1128 EU600913 clade_313 N NLactobacillus sp. KLDS 1.0711 1129 EU600915 clade_313 N N Lactobacillussp. KLDS 1.0713 1131 EU600917 clade_313 N N Lactobacillus sp. KLDS1.0716 1132 EU600921 clade_313 N N Lactobacillus sp. KLDS 1.0718 1133EU600922 clade_313 N N Lactobacillus sp. oral taxon 052 1137 GQ422710clade_313 N N Lactobacillus vaginalis 1140 ACGV01000168 clade_313 N NBrevibacterium aurantiacum 419 NR_044854 clade_314 N N Brevibacteriumlinens 423 AJ315491 clade_314 N N Lactobacillus pentosus 1108 JN813103clade_315 N N Lactobacillus plantarum 1110 ACGZ02000033 clade_315 N NLactobacillus sp. KLDS 1.0702 1123 EU600906 clade_315 N N Lactobacillussp. KLDS 1.0703 1124 EU600907 clade_315 N N Lactobacillus sp. KLDS1.0704 1125 EU600908 clade_315 N N Lactobacillus sp. KLDS 1.0705 1126EU600909 clade_315 N N Agrobacterium radiobacter 115 CP000628 clade_316N N Agrobacterium tumefaciens 116 AJ389893 clade_316 N N Corynebacteriumargentoratense 685 EF463055 clade_317 N N Corynebacterium diphtheriae693 NC_002935 clade_317 N OP Corynebacterium pseudotuberculosis 715NR_037070 clade_317 N N Corynebacterium renale 717 NR_037069 clade_317 NN Corynebacterium ulcerans 731 NR_074467 clade_317 N N Aurantimonascoralicida 191 AY065627 clade_318 N N Aureimonas altamirensis 192FN658986 clade_318 N N Lactobacillus acidipiscis 1066 NR_024718clade_320 N N Lactobacillus salivarius 1117 AEBA01000145 clade_320 N NLactobacillus sp. KLDS 1.0719 1134 EU600923 clade_320 N N Lactobacillusbuchneri 1073 ACGH01000101 clade_321 N N Lactobacillus genomosp. C1 1086AY278619 clade_321 N N Lactobacillus genomosp. C2 1087 AY278620clade_321 N N Lactobacillus hilgardii 1089 ACGP01000200 clade_321 N NLactobacillus kefiri 1096 NR_042230 clade_321 N N Lactobacillusparabuchneri 1105 NR_041294 clade_321 N N Lactobacillus parakefiri 1107NR_029039 clade_321 N N Lactobacillus curvatus 1079 NR_042437 clade_322N N Lactobacillus sakei 1116 DQ989236 clade_322 N N Aneurinibacillusaneurinilyticus 167 AB101592 clade_323 N N Aneurinibacillus danicus 168NR_028657 clade_323 N N Aneurinibacillus migulanus 169 NR_036799clade_323 N N Aneurinibacillus terranovensis 170 NR_042271 clade_323 N NStaphylococcus aureus 1757 CP002643 clade_325 N Category-BStaphylococcus auricularis 1758 JQ624774 clade_325 N N Staphylococcuscapitis 1759 ACFR01000029 clade_325 N N Staphylococcus caprae 1760ACRH01000033 clade_325 N N Staphylococcus carnosus 1761 NR_075003clade_325 N N Staphylococcus cohnii 1762 JN175375 clade_325 N NStaphylococcus condimenti 1763 NR_029345 clade_325 N N Staphylococcusepidermidis 1764 ACHE01000056 clade_325 N N Staphylococcus equorum 1765NR_027520 clade_325 N N Staphylococcus haemolyticus 1767 NC_007168clade_325 N N Staphylococcus hominis 1768 AM157418 clade_325 N NStaphylococcus lugdunensis 1769 AEQA01000024 clade_325 N NStaphylococcus pasteuri 1770 FJ189773 clade_325 N N Staphylococcuspseudintermedius 1771 CP002439 clade_325 N N Staphylococcussaccharolyticus 1772 NR_029158 clade_325 N N Staphylococcussaprophyticus 1773 NC_007350 clade_325 N N Staphylococcus sp. clonebottae7 1777 AF467424 clade_325 N N Staphylococcus sp. H292 1775AB177642 clade_325 N N Staphylococcus sp. H780 1776 AB177644 clade_325 NN Staphylococcus succinus 1778 NR_028667 clade_325 N N Staphylococcuswarneri 1780 ACPZ01000009 clade_325 N N Staphylococcus xylosus 1781AY395016 clade_325 N N Cardiobacterium hominis 490 ACKY01000036clade_326 N N Cardiobacterium valvarum 491 NR_028847 clade_326 N NPseudomonas fluorescens 1593 AY622220 clade_326 N N Pseudomonasgessardii 1594 FJ943496 clade_326 N N Pseudomonas monteilii 1596NR_024910 clade_326 N N Pseudomonas poae 1597 GU188951 clade_326 N NPseudomonas putida 1599 AF094741 clade_326 N N Pseudomonas sp. G12291601 DQ910482 clade_326 N N Pseudomonas tolaasii 1604 AF320988 clade_326N N Pseudomonas viridiflava 1605 NR_042764 clade_326 N N Listeria grayi1185 ACCR02000003 clade_328 N OP Listeria innocua 1186 JF967625clade_328 N N Listeria ivanovii 1187 X56151 clade_328 N N Listeriamonocytogenes 1188 CP002003 clade_328 N Category-B Listeria welshimeri1189 AM263198 clade_328 N OP Capnocytophaga sp. oral clone ASCH05 484AY923149 clade_333 N N Capnocytophaga sputigena 489 ABZV01000054clade_333 N N Leptotrichia genomosp. C1 1166 AY278621 clade_334 N NLeptotrichia shahii 1169 AY029806 clade_334 N N Leptotrichia sp.neutropenicPatient 1170 AF189244 clade_334 N N Leptotrichia sp. oralclone GT018 1171 AY349384 clade_334 N N Leptotrichia sp. oral cloneGT020 1172 AY349385 clade_334 N N Bacteroides sp. 20_3 296 ACRQ01000064clade_335 N N Bacteroides sp. 3_1_19 307 ADCJ01000062 clade_335 N NBacteroides sp. 3_2_5 311 ACIB01000079 clade_335 N N Parabacteroidesdistasonis 1416 CP000140 clade_335 N N Parabacteroides goldsteinii 1417AY974070 clade_335 N N Parabacteroides gordonii 1418 AB470344 clade_335N N Parabacteroides sp. D13 1421 ACPW01000017 clade_335 N NCapnocytophaga genomosp. C1 477 AY278613 clade_336 N N Capnocytophagaochracea 480 AEOH01000054 clade_336 N N Capnocytophaga sp. GEJ8 481GU561335 clade_336 N N Capnocytophaga sp. oral strain A47ROY 486AY005077 clade_336 N N Capnocytophaga sp. S1b 482 U42009 clade_336 N NParaprevotella clara 1426 AFFY01000068 clade_336 N N Bacteroidesheparinolyticus 282 JN867284 clade_338 N N Prevotella heparinolytica1500 GQ422742 clade_338 N N Treponema genomosp. P4 oral clone MB2_G191928 DQ003618 clade_339 N N Treponema genomosp. P6 oral clone MB4_G111930 DQ003625 clade_339 N N Treponema sp. oral taxon 254 1952 GU408803clade_339 N N Treponema sp. oral taxon 508 1956 GU413616 clade_339 N NTreponema sp. oral taxon 518 1957 GU413640 clade_339 N N Chlamydiamuridarum 502 AE002160 clade_341 N OP Chlamydia trachomatis 504 U68443clade_341 N OP Chlamydia psittaci 503 NR_036864 clade_342 N Category-BChlamydophila pneumoniae 509 NC_002179 clade_342 N OP Chlamydophilapsittaci 510 D85712 clade_342 N OP Anaerococcus octavius 146 NR_026360clade_343 N N Anaerococcus sp. 8405254 149 HM587319 clade_343 N NAnaerococcus sp. 9401487 150 HM587322 clade_343 N N Anaerococcus sp.9403502 151 HM587325 clade_343 N N Gardnerella vaginalis 923 CP001849clade_344 N N Campylobacter lari 466 CP000932 clade_346 N OPAnaerobiospirillum succiniciproducens 142 NR_026075 clade_347 N NAnaerobiospirillum thomasii 143 AJ420985 clade_347 N N Ruminobacteramylophilus 1654 NR_026450 clade_347 N N Succinatimonas hippei 1897AEVO01000027 clade_347 N N Actinomyces europaeus 54 NR_026363 clade_348N N Actinomyces sp. oral clone GU009 82 AY349361 clade_348 N N Moraxellacatarrhalis 1260 CP002005 clade_349 N N Moraxella lincolnii 1261FR822735 clade_349 N N Moraxella sp. 16285 1263 JF682466 clade_349 N NPsychrobacter sp. 13983 1613 HM212668 clade_349 N N Actinobaculummassiliae 49 AF487679 clade_350 N N Actinobaculum schaalii 50 AY957507clade_350 N N Actinobaculum sp. BM#101342 51 AY282578 clade_350 N NActinobaculum sp. P2P_19 P1 52 AY207066 clade_350 N N Actinomyces sp.oral clone IO076 84 AY349363 clade_350 N N Actinomyces sp. oral taxon848 93 ACUY01000072 clade_350 N N Actinomyces neuii 65 X71862 clade_352N N Mobiluncus mulieris 1252 ACKW01000035 clade_352 N N Blastomonasnatatoria 372 NR_040824 clade_356 N N Novosphingobium aromaticivorans1357 AAAV03000008 clade_356 N N Sphingomonas sp. oral clone FI012 1745AY349411 clade_356 N N Sphingopyxis alaskensis 1749 CP000356 clade_356 NN Oxalobacter formigenes 1389 ACDQ01000020 clade_357 N N Veillonellaatypica 1974 AEDS01000059 clade_358 N N Veillonella dispar 1975ACIK02000021 clade_358 N N Veillonella genomosp. P1 oral clone MB5_P171976 DQ003631 clade_358 N N Veillonella parvula 1978 ADFU01000009clade_358 N N Veillonella sp. 3_1_44 1979 ADCV01000019 clade_358 N NVeillonella sp. 6_1_27 1980 ADCW01000016 clade_358 N N Veillonella sp.ACP1 1981 HQ616359 clade_358 N N Veillonella sp. AS16 1982 HQ616365clade_358 N N Veillonella sp. BS32b 1983 HQ616368 clade_358 N NVeillonella sp. ICM51a 1984 HQ616396 clade_358 N N Veillonella sp. MSA121985 HQ616381 clade_358 N N Veillonella sp. NVG 100cf 1986 EF108443clade_358 N N Veillonella sp. OK11 1987 JN695650 clade_358 N NVeillonella sp. oral clone ASCG01 1990 AY923144 clade_358 N NVeillonella sp. oral clone ASCG02 1991 AY953257 clade_358 N NVeillonella sp. oral clone OH1A 1992 AY947495 clade_358 N N Veillonellasp. oral taxon 158 1993 AENU01000007 clade_358 N N Kocuria marina 1040GQ260086 clade_365 N N Kocuria rhizophila 1042 AY030315 clade_365 N NKocuria rosea 1043 X87756 clade_365 N N Kocuria varians 1044 AF542074clade_365 N N Clostridiaceae bacterium END_2 531 EF451053 clade_368 N NMicrococcus antarcticus 1242 NR_025285 clade_371 N N Micrococcus luteus1243 NR_075062 clade_371 N N Micrococcus lylae 1244 NR_026200 clade_371N N Micrococcus sp. 185 1245 EU714334 clade_371 N N Lactobacillus brevis1072 EU194349 clade_372 N N Lactobacillus parabrevis 1104 NR_042456clade_372 N N Pediococcus acidilactici 1436 ACXB01000026 clade_372 N NPediococcus pentosaceus 1437 NR_075052 clade_372 N N Lactobacillusdextrinicus 1081 NR_036861 clade_373 N N Lactobacillus perolens 1109NR_029360 clade_373 N N Lactobacillus rhamnosus 1113 ABWJ01000068clade_373 N N Lactobacillus saniviri 1118 AB602569 clade_373 N NLactobacillus sp. BT6 1121 HQ616370 clade_373 N N Mycobacteriummageritense 1282 FR798914 clade_374 N OP Mycobacterium neoaurum 1286AF268445 clade_374 N OP Mycobacterium smegmatis 1291 CP000480 clade_374N OP Mycobacterium sp. HE5 1304 AJ012738 clade_374 N N Dysgonomonasgadei 775 ADLV01000001 clade_377 N N Dysgonomonas mossii 776ADLW01000023 clade_377 N N Porphyromonas levii 1474 NR_025907 clade_377N N Porphyromonas somerae 1476 AB547667 clade_377 N N Bacteroidesbarnesiae 267 NR_041446 clade_378 N N Bacteroides coprocola 272ABIY02000050 clade_378 N N Bacteroides coprophilus 273 ACBW01000012clade_378 N N Bacteroides dorei 274 ABWZ01000093 clade_378 N NBacteroides massiliensis 284 AB200226 clade_378 N N Bacteroides plebeius289 AB200218 clade_378 N N Bacteroides sp. 3_1_33FAA 309 ACPS01000085clade_378 N N Bacteroides sp. 3_1_40A 310 ACRT01000136 clade_378 N NBacteroides sp. 4_3_47FAA 313 ACDR02000029 clade_378 N N Bacteroides sp.9_1_42FAA 314 ACAA01000096 clade_378 N N Bacteroides sp. NB_8 323AB117565 clade_378 N N Bacteroides vulgatus 331 CP000139 clade_378 N NBacteroides ovatus 287 ACWH01000036 clade_38 N N Bacteroides sp. 1_1_30294 ADCL01000128 clade_38 N N Bacteroides sp. 2_1_22 297 ACPQ01000117clade_38 N N Bacteroides sp. 2_2_4 299 ABZZ01000168 clade_38 N NBacteroides sp. 3_1_23 308 ACRS01000081 clade_38 N N Bacteroides sp. D1318 ACAB02000030 clade_38 N N Bacteroides sp. D2 321 ACGA01000077clade_38 N N Bacteroides sp. D22 320 ADCK01000151 clade_38 N NBacteroides xylanisolvens 332 ADKP01000087 clade_38 N N Treponemalecithinolyticum 1931 NR_026247 clade_380 N OP Treponema parvum 1933AF302937 clade_380 N OP Treponema sp. oral clone JU025 1940 AY349417clade_380 N N Treponema sp. oral taxon 270 1954 GQ422733 clade_380 N NParascardovia denticolens 1428 ADEB01000020 clade_381 N N Scardoviainopinata 1688 AB029087 clade_381 N N Scardovia wiggsiae 1689 AY278626clade_381 N N Clostridiales bacterium 9400853 533 HM587320 clade_384 N NMogibacterium diversum 1254 NR_027191 clade_384 N N Mogibacteriumneglectum 1255 NR_027203 clade_384 N N Mogibacterium pumilum 1256NR_028608 clade_384 N N Mogibacterium timidum 1257 Z36296 clade_384 N NBorrelia burgdorferi 389 ABGI01000001 clade_386 N OP Borrelia garinii392 ABJV01000001 clade_386 N OP Borrelia sp. NE49 397 AJ224142 clade_386N OP Caldimonas manganoxidans 457 NR_040787 clade_387 N N Comamonadaceaebacterium oral taxon F47 667 HM099651 clade_387 N N Lautropia mirabilis1149 AEQP01000026 clade_387 N N Lautropia sp. oral clone AP009 1150AY005030 clade_387 N N Peptoniphilus asaccharolyticus 1441 D14145clade_389 N N Peptoniphilus duerdenii 1442 EU526290 clade_389 N NPeptoniphilus harei 1443 NR_026358 clade_389 N N Peptoniphilus indolicus1444 AY153431 clade_389 N N Peptoniphilus lacrimalis 1446 ADDO01000050clade_389 N N Peptoniphilus sp. gpac077 1450 AM176527 clade_389 N NPeptoniphilus sp. JC140 1447 JF824803 clade_389 N N Peptoniphilus sp.oral taxon 386 1452 ADCS01000031 clade_389 N N Peptoniphilus sp. oraltaxon 836 1453 AEAA01000090 clade_389 N N Peptostreptococcaceaebacterium ph1 1454 JN837495 clade_389 N N Dialister pneumosintes 765HM596297 clade_390 N N Dialister sp. oral taxon 502 767 GQ422739clade_390 N N Cupriavidus metallidurans 741 GU230889 clade_391 N NHerbaspirillum seropedicae 1001 CP002039 clade_391 N N Herbaspirillumsp. JC206 1002 JN657219 clade_391 N N Janthinobacterium sp. SY12 1015EF455530 clade_391 N N Massilia sp. CCUG 43427A 1197 FR773700 clade_391N N Ralstonia pickettii 1615 NC_010682 clade_391 N N Ralstonia sp.5_7_47FAA 1616 ACUF01000076 clade_391 N N Francisella novicida 889ABSS01000002 clade_392 N N Francisella philomiragia 890 AY928394clade_392 N N Francisella tularensis 891 ABAZ01000082 clade_392 NCategory-A Ignatzschineria indica 1009 HQ823562 clade_392 N NIgnatzschineria sp. NML 95_0260 1010 HQ823559 clade_392 N NStreptococcus mutans 1814 AP010655 clade_394 N N Lactobacillus gasseri1084 ACOZ01000018 clade_398 N N Lactobacillus hominis 1090 FR681902clade_398 N N Lactobacillus iners 1091 AEKJ01000002 clade_398 N NLactobacillus johnsonii 1093 AE017198 clade_398 N N Lactobacillussenioris 1119 AB602570 clade_398 N N Lactobacillus sp. oral clone HT0021135 AY349382 clade_398 N N Weissella beninensis 2006 EU439435 clade_398N N Sphingomonas echinoides 1744 NR_024700 clade_399 N N Sphingomonassp. oral taxon A09 1747 HM099639 clade_399 N N Sphingomonas sp. oraltaxon F71 1748 HM099645 clade_399 N N Zymomonas mobilis 2032 NR_074274clade_399 N N Arcanobacterium haemolyticum 174 NR_025347 clade_400 N NArcanobacterium pyogenes 175 GU585578 clade_400 N N Trueperella pyogenes1962 NR_044858 clade_400 N N Lactococcus garvieae 1144 AF061005clade_401 N N Lactococcus lactis 1145 CP002365 clade_401 N NBrevibacterium mcbrellneri 424 ADNU01000076 clade_402 N N Brevibacteriumpaucivorans 425 EU086796 clade_402 N N Brevibacterium sp. JC43 428JF824806 clade_402 N N Selenomonas artemidis 1692 HM596274 clade_403 N NSelenomonas sp. FOBRC9 1704 HQ616378 clade_403 N N Selenomonas sp. oraltaxon 137 1715 AENV01000007 clade_403 N N Desmospora activa 751 AM940019clade_404 N N Desmospora sp. 8437 752 AFHT01000143 clade_404 N NPaenibacillus sp. oral taxon F45 1407 HM099647 clade_404 N NCorynebacterium ammoniagenes 682 ADNS01000011 clade_405 N NCorynebacterium aurimucosum 687 ACLH01000041 clade_405 N NCorynebacterium bovis 688 AF537590 clade_405 N N Corynebacterium canis689 GQ871934 clade_405 N N Corynebacterium casei 690 NR_025101 clade_405N N Corynebacterium durum 694 Z97069 clade_405 N N Corynebacteriumefficiens 695 ACLI01000121 clade_405 N N Corynebacterium falsenii 696Y13024 clade_405 N N Corynebacterium flavescens 697 NR_037040 clade_405N N Corynebacterium glutamicum 701 BA000036 clade_405 N NCorynebacterium jeikeium 704 ACYW01000001 clade_405 N OP Corynebacteriumkroppenstedtii 705 NR_026380 clade_405 N N Corynebacteriumlipophiloflavum 706 ACHJ01000075 clade_405 N N Corynebacteriummatruchotii 709 ACSH02000003 clade_405 N N Corynebacterium minutissimum710 X82064 clade_405 N N Corynebacterium resistens 718 ADGN01000058clade_405 N N Corynebacterium simulans 720 AF537604 clade_405 N NCorynebacterium singulare 721 NR_026394 clade_405 N N Corynebacteriumsp. 1 ex sheep 722 Y13427 clade_405 N N Corynebacterium sp. NML 99_0018726 GU238413 clade_405 N N Corynebacterium striatum 727 ACGE01000001clade_405 N OP Corynebacterium urealyticum 732 X81913 clade_405 N OPCorynebacterium variabile 734 NR_025314 clade_405 N N Aerococcussanguinicola 98 AY837833 clade_407 N N Aerococcus urinae 99 CP002512clade_407 N N Aerococcus urinaeequi 100 NR_043443 clade_407 N NAerococcus viridans 101 ADNT01000041 clade_407 N N Fusobacteriumnaviforme 898 HQ223106 clade_408 N N Moryella indoligenes 1268 AF527773clade_408 N N Selenomonas genomosp. P5 1697 AY341820 clade_410 N NSelenomonas sp. oral clone IQ048 1710 AY349408 clade_410 N N Selenomonassputigena 1717 ACKP02000033 clade_410 N N Hyphomicrobium sulfonivorans1007 AY468372 clade_411 N N Methylocella silvestris 1228 NR_074237clade_411 N N Legionella pneumophila 1153 NC_002942 clade_412 N OPLactobacillus coryniformis 1077 NR_044705 clade_413 N N Arthrobacteragilis 178 NR_026198 clade_414 N N Arthrobacter arilaitensis 179NR_074608 clade_414 N N Arthrobacter bergerei 180 NR_025612 clade_414 NN Arthrobacter globiformis 181 NR_026187 clade_414 N N Arthrobacternicotianae 182 NR_026190 clade_414 N N Mycobacterium abscessus 1269AGQU01000002 clade_418 N OP Mycobacterium chelonae 1273 AB548610clade_418 N OP Bacteroides salanitronis 291 CP002530 clade_419 N NParaprevotella xylaniphila 1427 AFBR01000011 clade_419 N N Barnesiellaintestinihominis 336 AB370251 clade_420 N N Barnesiella viscericola 337NR_041508 clade_420 N N Parabacteroides sp. NS31_3 1422 JN029805clade_420 N N Porphyromonadaceae bacterium NML 060648 1470 EF184292clade_420 N N Tannerella forsythia 1913 CP003191 clade_420 N NTannerella sp. 6_1_58FAA_CT1 1914 ACWX01000068 clade_420 N N Mycoplasmaamphoriforme 1311 AY531656 clade_421 N N Mycoplasma genitalium 1317L43967 clade_421 N N Mycoplasma pneumoniae 1322 NC_000912 clade_421 N NMycoplasma penetrans 1321 NC_004432 clade_422 N N Ureaplasma parvum 1966AE002127 clade_422 N N Ureaplasma urealyticum 1967 AAYN01000002clade_422 N N Treponema genomosp. P1 1927 AY341822 clade_425 N NTreponema sp. oral taxon 228 1943 GU408580 clade_425 N N Treponema sp.oral taxon 230 1944 GU408603 clade_425 N N Treponema sp. oral taxon 2311945 GU408631 clade_425 N N Treponema sp. oral taxon 232 1946 GU408646clade_425 N N Treponema sp. oral taxon 235 1947 GU408673 clade_425 N NTreponema sp. ovine footrot 1959 AJ010951 clade_425 N N Treponemavincentii 1960 ACYH01000036 clade_425 N OP Burkholderiales bacterium1_1_47 452 ADCQ01000066 clade_432 N OP Parasutterella excrementihominis1429 AFBP01000029 clade_432 N N Parasutterella secunda 1430 AB491209clade_432 N N Sutterella morbirenis 1898 AJ832129 clade_432 N NSutterella sanguinus 1900 AJ748647 clade_432 N N Sutterella sp. YIT12072 1901 AB491210 clade_432 N N Sutterella stercoricanis 1902NR_025600 clade_432 N N Sutterella wadsworthensis 1903 ADMF01000048clade_432 N N Propionibacterium freudenreichii 1572 NR_036972 clade_433N N Propionibacterium sp. oral taxon 192 1580 GQ422728 clade_433 N NTessaracoccus sp. oral taxon F04 1917 HM099640 clade_433 N NPeptoniphilus ivorii 1445 Y07840 clade_434 N N Peptoniphilus sp. gpac0071448 AM176517 clade_434 N N Peptoniphilus sp. gpac018A 1449 AM176519clade_434 N N Peptoniphilus sp. gpac148 1451 AM176535 clade_434 N NFlexispira rappini 887 AY126479 clade_436 N N Helicobacter bilis 993ACDN01000023 clade_436 N N Helicobacter cinaedi 995 ABQT01000054clade_436 N N Helicobacter sp. None 998 U44756 clade_436 N NBrevundimonas subvibrioides 429 CP002102 clade_438 N N Hyphomonasneptunium 1008 NR_074092 clade_438 N N Phenylobacterium zucineum 1465AY628697 clade_438 N N Streptococcus downei 1793 AEKN01000002 clade_441N N Streptococcus sp. SHV515 1848 Y07601 clade_441 N N Acinetobacter sp.CIP 53.82 40 JQ638584 clade_443 N N Halomonas elongata 990 NR_074782clade_443 N N Halomonas johnsoniae 991 FR775979 clade_443 N NButyrivibrio fibrisolvens 456 U41172 clade_444 N N Roseburia sp. 11SE371640 FM954975 clade_444 N N Roseburia sp. 11SE38 1641 FM954976 clade_444N N Shuttleworthia satelles 1728 ACIP02000004 clade_444 N NShuttleworthia sp. MSX8B 1729 HQ616383 clade_444 N N Shuttleworthia sp.oral taxon G69 1730 GU432167 clade_444 N N Bdellovibrio sp. MPA 344AY294215 clade_445 N N Desulfobulbus sp. oral clone CH031 755 AY005036clade_445 N N Desulfovibrio desulfuricans 757 DQ092636 clade_445 N NDesulfovibrio fairfieldensis 758 U42221 clade_445 N N Desulfovibriopiger 759 AF192152 clade_445 N N Desulfovibrio sp. 3_1_syn3 760ADDR01000239 clade_445 N N Geobacter bemidjiensis 941 CP001124 clade_445N N Brachybacterium alimentarium 401 NR_026269 clade_446 N NBrachybacterium conglomeratum 402 AB537169 clade_446 N N Brachybacteriumtyrofermentans 403 NR_026272 clade_446 N N Dermabacter hominis 749FJ263375 clade_446 N N Aneurinibacillus thermoaerophilus 171 NR_029303clade_448 N N Brevibacillus agri 409 NR_040983 clade_448 N NBrevibacillus centrosporus 411 NR_043414 clade_448 N N Brevibacilluschoshinensis 412 NR_040980 clade_448 N N Brevibacillus invocatus 413NR_041836 clade_448 N N Brevibacillus parabrevis 415 NR_040981 clade_448N N Brevibacillus reuszeri 416 NR_040982 clade_448 N N Brevibacillus sp.phR 417 JN837488 clade_448 N N Brevibacillus thermoruber 418 NR_026514clade_448 N N Lactobacillus murinus 1100 NR_042231 clade_449 N NLactobacillus oeni 1102 NR_043095 clade_449 N N Lactobacillus ruminis1115 ACGS02000043 clade_449 N N Lactobacillus vini 1141 NR_042196clade_449 N N Gemella haemolysans 924 ACDZ02000012 clade_450 N N Gemellamorbillorum 925 NR_025904 clade_450 N N Gemella morbillorum 926ACRX01000010 clade_450 N N Gemella sanguinis 927 ACRY01000057 clade_450N N Gemella sp. oral clone ASCE02 929 AY923133 clade_450 N N Gemella sp.oral clone ASCF04 930 AY923139 clade_450 N N Gemella sp. oral cloneASCF12 931 AY923143 clade_450 N N Gemella sp. WAL 1945J 928 EU427463clade_450 N N Sporolactobacillus nakayamae 1753 NR_042247 clade_451 N NGluconacetobacter entanii 945 NR_028909 clade_452 N N Gluconacetobactereuropaeus 946 NR_026513 clade_452 N N Gluconacetobacter hansenii 947NR_026133 clade_452 N N Gluconacetobacter oboediens 949 NR_041295clade_452 N N Gluconacetobacter xylinus 950 NR_074338 clade_452 N NAuritibacter ignavus 193 FN554542 clade_453 N N Dermacoccus sp. Ellin185750 AEIQ01000090 clade_453 N N Janibacter limosus 1013 NR_026362clade_453 N N Janibacter melonis 1014 EF063716 clade_453 N N Acetobacteraceti 7 NR_026121 clade_454 N N Acetobacter fabarum 8 NR_042678clade_454 N N Acetobacter lovaniensis 9 NR_040832 clade_454 N NAcetobacter malorum 10 NR_025513 clade_454 N N Acetobacter orientalis 11NR_028625 clade_454 N N Acetobacter pasteurianus 12 NR_026107 clade_454N N Acetobacter pomorum 13 NR_042112 clade_454 N N Acetobacter syzygii14 NR_040868 clade_454 N N Acetobacter tropicalis 15 NR_036881 clade_454N N Gluconacetobacter azotocaptans 943 NR_028767 clade_454 N NGluconacetobacter diazotrophicus 944 NR_074292 clade_454 N NGluconacetobacter johannae 948 NR_024959 clade_454 N N Nocardiabrasiliensis 1351 AIHV01000038 clade_455 N N Nocardia cyriacigeorgica1352 HQ009486 clade_455 N N Nocardia puris 1354 NR_028994 clade_455 N NNocardia sp. 01_Je_025 1355 GU574059 clade_455 N N Rhodococcus equi 1623ADNW01000058 clade_455 N N Oceanobacillus caeni 1358 NR_041533 clade_456N N Oceanobacillus sp. Ndiop 1359 CAER01000083 clade_456 N NOrnithinibacillus bavariensis 1384 NR_044923 clade_456 N NOrnithinibacillus sp. 7_10AIA 1385 FN397526 clade_456 N N Virgibacillusproomii 2005 NR_025308 clade_456 N N Corynebacterium amycolatum 683ABZU01000033 clade_457 N OP Corynebacterium hansenii 702 AM946639clade_457 N N Corynebacterium xerosis 735 FN179330 clade_457 N OPStaphylococcaceae bacterium NML 92_0017 1756 AY841362 clade_458 N NStaphylococcus fleurettii 1766 NR_041326 clade_458 N N Staphylococcussciuri 1774 NR_025520 clade_458 N N Staphylococcus vitulinus 1779NR_024670 clade_458 N N Stenotrophomonas maltophilia 1782 AAVZ01000005clade_459 N N Stenotrophomonas sp. FG_6 1783 EF017810 clade_459 N NMycobacterium africanum 1270 AF480605 clade_46 N OP Mycobacteriumalsiensis 1271 AJ938169 clade_46 N OP Mycobacterium avium 1272 CP000479clade_46 N OP Mycobacterium colombiense 1274 AM062764 clade_46 N OPMycobacterium gordonae 1276 GU142930 clade_46 N OP Mycobacteriumintracellulare 1277 GQ153276 clade_46 N OP Mycobacterium kansasii 1278AF480601 clade_46 N OP Mycobacterium lacus 1279 NR_025175 clade_46 N OPMycobacterium leprae 1280 FM211192 clade_46 N OP Mycobacteriumlepromatosis 1281 EU203590 clade_46 N OP Mycobacterium mantenii 1283FJ042897 clade_46 N OP Mycobacterium marinum 1284 NC_010612 clade_46 NOP Mycobacterium microti 1285 NR_025234 clade_46 N OP Mycobacteriumparascrofulaceum 1287 ADNV01000350 clade_46 N OP Mycobacterium seoulense1290 DQ536403 clade_46 N OP Mycobacterium sp. 1761 1292 EU703150clade_46 N N Mycobacterium sp. 1791 1295 EU703148 clade_46 N NMycobacterium sp. 1797 1296 EU703149 clade_46 N N Mycobacterium sp.B10_07.09.0206 1298 HQ174245 clade_46 N N Mycobacterium sp. NLA0010007361305 HM627011 clade_46 N N Mycobacterium sp. W 1306 DQ437715 clade_46 NN Mycobacterium tuberculosis 1307 CP001658 clade_46 N Category-CMycobacterium ulcerans 1308 AB548725 clade_46 N OP Mycobacteriumvulneris 1309 EU834055 clade_46 N OP Xanthomonas campestris 2016EF101975 clade_461 N N Xanthomonas sp. kmd_489 2017 EU723184 clade_461 NN Dietzia natronolimnaea 769 GQ870426 clade_462 N N Dietzia sp. BBDP51770 DQ337512 clade_462 N N Dietzia sp. CA149 771 GQ870422 clade_462 N NDietzia timorensis 772 GQ870424 clade_462 N N Gordonia bronchialis 951NR_027594 clade_463 N N Gordonia polyisoprenivorans 952 DQ385609clade_463 N N Gordonia sp. KTR9 953 DQ068383 clade_463 N N Gordoniasputi 954 FJ536304 clade_463 N N Gordonia terrae 955 GQ848239 clade_463N N Leptotrichia goodfellowii 1167 ADAD01000110 clade_465 N NLeptotrichia sp. oral clone IK040 1174 AY349387 clade_465 N NLeptotrichia sp. oral clone P2PB_51 P1 1175 AY207053 clade_465 N NBacteroidales genomosp. P7 oral clone MB3_P19 264 DQ003623 clade_466 N NButyricimonas virosa 454 AB443949 clade_466 N N Odoribacter laneus 1363AB490805 clade_466 N N Odoribacter splanchnicus 1364 CP002544 clade_466N N Capnocytophaga gingivalis 478 ACLQ01000011 clade_467 N NCapnocytophaga granulosa 479 X97248 clade_467 N N Capnocytophaga sp.oral clone AH015 483 AY005074 clade_467 N N Capnocytophaga sp. oralstrain S3 487 AY005073 clade_467 N N Capnocytophaga sp. oral taxon 338488 AEXX01000050 clade_467 N N Capnocytophaga canimorsus 476 CP002113clade_468 N N Capnocytophaga sp. oral clone ID062 485 AY349368 clade_468N N Lactobacillus catenaformis 1075 M23729 clade_469 N N Lactobacillusvitulinus 1142 NR_041305 clade_469 N N Cetobacterium somerae 501AJ438155 clade_470 N N Fusobacterium gonidiaformans 896 ACET01000043clade_470 N N Fusobacterium mortiferum 897 ACDB02000034 clade_470 N NFusobacterium necrogenes 899 X55408 clade_470 N N Fusobacteriumnecrophorum 900 AM905356 clade_470 N N Fusobacterium sp. 12_1B 905AGWJ01000070 clade_470 N N Fusobacterium sp. 3_1_5R 911 ACDD01000078clade_470 N N Fusobacterium sp. D12 918 ACDG02000036 clade_470 N NFusobacterium ulcerans 921 ACDH01000090 clade_470 N N Fusobacteriumvarium 922 ACIE01000009 clade_470 N N Mycoplasma arthritidis 1312NC_011025 clade_473 N N Mycoplasma faucium 1314 NR_024983 clade_473 N NMycoplasma hominis 1318 AF443616 clade_473 N N Mycoplasma orale 1319AY796060 clade_473 N N Mycoplasma salivarium 1324 M24661 clade_473 N NMitsuokella jalaludinii 1247 NR_028840 clade_474 N N Mitsuokellamultacida 1248 ABWK02000005 clade_474 N N Mitsuokella sp. oral taxon 5211249 GU413658 clade_474 N N Mitsuokella sp. oral taxon G68 1250 GU432166clade_474 N N Selenomonas genomosp. C1 1695 AY278627 clade_474 N NSelenomonas genomosp. P8 oral clone MB5_P06 1700 DQ003628 clade_474 N NSelenomonas ruminantium 1703 NR_075026 clade_474 N N Veillonellaceaebacterium oral taxon 131 1994 GU402916 clade_474 N N Alloscardoviaomnicolens 139 NR_042583 clade_475 N N Alloscardovia sp. OB7196 140AB425070 clade_475 N N Bifidobacterium urinalis 366 AJ278695 clade_475 NN Prevotella loescheii 1503 JN867231 clade_48 N N Prevotella sp. oralclone ASCG12 1530 DQ272511 clade_48 N N Prevotella sp. oral clone GU0271540 AY349398 clade_48 N N Prevotella sp. oral taxon 472 1553ACZS01000106 clade_48 N N Selenomonas dianae 1693 GQ422719 clade_480 N NSelenomonas flueggei 1694 AF287803 clade_480 N N Selenomonas genomosp.C2 1696 AY278628 clade_480 N N Selenomonas genomosp. P6 oral cloneMB3_C41 1698 DQ003636 clade_480 N N Selenomonas genomosp. P7 oral cloneMB5_C08 1699 DQ003627 clade_480 N N Selenomonas infelix 1701 AF287802clade_480 N N Selenomonas noxia 1702 GU470909 clade_480 N N Selenomonassp. oral clone FT050 1705 AY349403 clade_480 N N Selenomonas sp. oralclone GI064 1706 AY349404 clade_480 N N Selenomonas sp. oral clone GT0101707 AY349405 clade_480 N N Selenomonas sp. oral clone HU051 1708AY349406 clade_480 N N Selenomonas sp. oral clone IK004 1709 AY349407clade_480 N N Selenomonas sp. oral clone JI021 1711 AY349409 clade_480 NN Selenomonas sp. oral clone JS031 1712 AY349410 clade_480 N NSelenomonas sp. oral clone OH4A 1713 AY947498 clade_480 N N Selenomonassp. oral clone P2PA_80 P4 1714 AY207052 clade_480 N N Selenomonas sp.oral taxon 149 1716 AEEJ01000007 clade_480 N N Veillonellaceae bacteriumoral taxon 155 1995 GU470897 clade_480 N N Agrococcus jenensis 117NR_026275 clade_484 N N Microbacterium gubbeenense 1232 NR_025098clade_484 N N Pseudoclavibacter sp. Timone 1590 FJ375951 clade_484 N NTropheryma whipplei 1961 BX251412 clade_484 N N Zimmermannella bifida2031 AB012592 clade_484 N N Legionella hackeliae 1151 M36028 clade_486 NOP Legionella longbeachae 1152 M36029 clade_486 N OP Legionella sp.D3923 1154 JN380999 clade_486 N OP Legionella sp. D4088 1155 JN381012clade_486 N OP Legionella sp. H63 1156 JF831047 clade_486 N OPLegionella sp. NML 93L054 1157 GU062706 clade_486 N OP Legionellasteelei 1158 HQ398202 clade_486 N OP Tatlockia micdadei 1915 M36032clade_486 N N Helicobacter pullorum 996 ABQU01000097 clade_489 N NAcetobacteraceae bacterium AT_5844 16 AGEZ01000040 clade_490 N NRoseomonas cervicalis 1643 ADVL01000363 clade_490 N N Roseomonas mucosa1644 NR_028857 clade_490 N N Roseomonas sp. NML94_0193 1645 AF533357clade_490 N N Roseomonas sp. NML97_0121 1646 AF533359 clade_490 N NRoseomonas sp. NML98_0009 1647 AF533358 clade_490 N N Roseomonas sp.NML98_0157 1648 AF533360 clade_490 N N Rickettsia akari 1627 CP000847clade_492 N OP Rickettsia conorii 1628 AE008647 clade_492 N OPRickettsia prowazekii 1629 M21789 clade_492 N Category-B Rickettsiarickettsii 1630 NC_010263 clade_492 N OP Rickettsia slovaca 1631 L36224clade_492 N OP Rickettsia typhi 1632 AE017197 clade_492 N OPAnaeroglobus geminatus 160 AGCJ01000054 clade_493 N N Megasphaeragenomosp. C1 1201 AY278622 clade_493 N N Megasphaera micronuciformis1203 AECS01000020 clade_493 N N Clostridiales genomosp. BVAB3 540CP001850 clade_495 N N Tsukamurella paurometabola 1963 X80628 clade_496N N Tsukamurella tyrosinosolvens 1964 AB478958 clade_496 N N Abiotrophiapara_adiacens 2 AB022027 clade_497 N N Carnobacterium divergens 492NR_044706 clade_497 N N Carnobacterium maltaromaticum 493 NC_019425clade_497 N N Enterococcus avium 800 AF133535 clade_497 N N Enterococcuscaccae 801 AY943820 clade_497 N N Enterococcus casseliflavus 802AEWT01000047 clade_497 N N Enterococcus durans 803 AJ276354 clade_497 NN Enterococcus faecalis 804 AE016830 clade_497 N N Enterococcus faecium805 AM157434 clade_497 N N Enterococcus gallinarum 806 AB269767clade_497 N N Enterococcus gilvus 807 AY033814 clade_497 N NEnterococcus hawaiiensis 808 AY321377 clade_497 N N Enterococcus hirae809 AF061011 clade_497 N N Enterococcus italicus 810 AEPV01000109clade_497 N N Enterococcus mundtii 811 NR_024906 clade_497 N NEnterococcus raffinosus 812 FN600541 clade_497 N N Enterococcus sp.BV2CASA2 813 JN809766 clade_497 N N Enterococcus sp. CCRI_16620 814GU457263 clade_497 N N Enterococcus sp. F95 815 FJ463817 clade_497 N NEnterococcus sp. RfL6 816 AJ133478 clade_497 N N Enterococcusthailandicus 817 AY321376 clade_497 N N Fusobacterium canifelinum 893AY162222 clade_497 N N Fusobacterium genomosp. C1 894 AY278616 clade_497N N Fusobacterium genomosp. C2 895 AY278617 clade_497 N N Fusobacteriumperiodonticum 902 ACJY01000002 clade_497 N N Fusobacterium sp. 1_1_41FAA906 ADGG01000053 clade_497 N N Fusobacterium sp. 11_3_2 904 ACUO01000052clade_497 N N Fusobacterium sp. 2_1_31 907 ACDC02000018 clade_497 N NFusobacterium sp. 3_1_27 908 ADGF01000045 clade_497 N N Fusobacteriumsp. 3_1_33 909 ACQE01000178 clade_497 N N Fusobacterium sp. 3_1_36A2 910ACPU01000044 clade_497 N N Fusobacterium sp. AC18 912 HQ616357 clade_497N N Fusobacterium sp. ACB2 913 HQ616358 clade_497 N N Fusobacterium sp.AS2 914 HQ616361 clade_497 N N Fusobacterium sp. CM1 915 HQ616371clade_497 N N Fusobacterium sp. CM21 916 HQ616375 clade_497 N NFusobacterium sp. CM22 917 HQ616376 clade_497 N N Fusobacterium sp. oralclone ASCF06 919 AY923141 clade_497 N N Fusobacterium sp. oral cloneASCF11 920 AY953256 clade_497 N N Granulicatella adiacens 959ACKZ01000002 clade_497 N N Granulicatella elegans 960 AB252689 clade_497N N Granulicatella paradiacens 961 AY879298 clade_497 N N Granulicatellasp. oral clone ASC02 963 AY923126 clade_497 N N Granulicatella sp. oralclone ASCA05 964 DQ341469 clade_497 N N Granulicatella sp. oral cloneASCB09 965 AY953251 clade_497 N N Granulicatella sp. oral clone ASCG05966 AY923146 clade_497 N N Tetragenococcus halophilus 1918 NR_075020clade_497 N N Tetragenococcus koreensis 1919 NR_043113 clade_497 N NVagococcus fluvialis 1973 NR_026489 clade_497 N N Chryseobacteriumanthropi 514 AM982793 clade_498 N N Chryseobacterium gleum 515ACKQ02000003 clade_498 N N Chryseobacterium hominis 516 NR_042517clade_498 N N Treponema refringens 1936 AF426101 clade_499 N OPTreponema sp. oral clone JU031 1941 AY349416 clade_499 N N Treponema sp.oral taxon 239 1948 GU408738 clade_499 N N Treponema sp. oral taxon 2711955 GU408871 clade_499 N N Alistipes finegoldii 129 NR_043064 clade_500N N Alistipes onderdonkii 131 NR_043318 clade_500 N N Alistipesputredinis 132 ABFK02000017 clade_500 N N Alistipes shahii 133 FP929032clade_500 N N Alistipes sp. HGB5 134 AENZ01000082 clade_500 N NAlistipes sp. JC50 135 JF824804 clade_500 N N Alistipes sp. RMA 9912 136GQ140629 clade_500 N N Mycoplasma agalactiae 1310 AF010477 clade_501 N NMycoplasma bovoculi 1313 NR_025987 clade_501 N N Mycoplasma fermentans1315 CP002458 clade_501 N N Mycoplasma flocculare 1316 X62699 clade_501N N Mycoplasma ovipneumoniae 1320 NR_025989 clade_501 N N Arcobacterbutzleri 176 AEPT01000071 clade_502 N N Arcobacter cryaerophilus 177NR_025905 clade_502 N N Campylobacter curvus 461 NC_009715 clade_502 NOP Campylobacter rectus 467 ACFU01000050 clade_502 N OP Campylobactershowae 468 ACVQ01000030 clade_502 N OP Campylobacter sp. FOBRC14 469HQ616379 clade_502 N OP Campylobacter sp. FOBRC15 470 HQ616380 clade_502N OP Campylobacter sp. oral clone BB120 471 AY005038 clade_502 N OPCampylobacter sputorum 472 NR_044839 clade_502 N OP Bacteroidesureolyticus 330 GQ167666 clade_504 N N Campylobacter gracilis 463ACYG01000026 clade_504 N OP Campylobacter hominis 464 NC_009714clade_504 N OP Dialister invisus 762 ACIM02000001 clade_506 N NDialister micraerophilus 763 AFBB01000028 clade_506 N N Dialistermicroaerophilus 764 AENT01000008 clade_506 N N Dialisterpropionicifaciens 766 NR_043231 clade_506 N N Dialister succinatiphilus768 AB370249 clade_506 N N Megasphaera elsdenii 1200 AY038996 clade_506N N Megasphaera genomosp. type_1 1202 ADGP01000010 clade_506 N NMegasphaera sp. BLPYG_07 1204 HM990964 clade_506 N N Megasphaera sp.UPII 199_6 1205 AFIJ01000040 clade_506 N N Chromobacterium violaceum 513NC_005085 clade_507 N N Laribacter hongkongensis 1148 CP001154 clade_507N N Methylophilus sp. ECd5 1229 AY436794 clade_507 N N Finegoldia magna883 ACHM02000001 clade_509 N N Parvimonas micra 1431 AB729072 clade_509N N Parvimonas sp. oral taxon 110 1432 AFII01000002 clade_509 N NPeptostreptococcus micros 1456 AM176538 clade_509 N N Peptostreptococcussp. oral clone FJ023 1460 AY349390 clade_509 N N Peptostreptococcus sp.P4P_31 P3 1458 AY207059 clade_509 N N Helicobacter pylori 997 CP000012clade_510 N OP Anaplasma marginale 165 ABOR01000019 clade_511 N NAnaplasma phagocytophilum 166 NC_007797 clade_511 N N Ehrlichiachaffeensis 783 AAIF01000035 clade_511 N OP Neorickettsia risticii 1349CP001431 clade_511 N N Neorickettsia sennetsu 1350 NC_007798 clade_511 NN Pseudoramibacter alactolyticus 1606 AB036759 clade_512 N N Veillonellamontpellierensis 1977 AF473836 clade_513 N N Veillonella sp. oral cloneASCA08 1988 AY923118 clade_513 N N Veillonella sp. oral clone ASCB031989 AY923122 clade_513 N N Inquilinus limosus 1012 NR_029046 clade_514N N Sphingomonas sp. oral clone FZ016 1746 AY349412 clade_514 N NAnaerococcus lactolyticus 145 ABYO01000217 clade_515 N N Anaerococcusprevotii 147 CP001708 clade_515 N N Anaerococcus sp. gpac104 152AM176528 clade_515 N N Anaerococcus sp. gpac126 153 AM176530 clade_515 NN Anaerococcus sp. gpac155 154 AM176536 clade_515 N N Anaerococcus sp.gpac199 155 AM176539 clade_515 N N Anaerococcus tetradius 157ACGC01000107 clade_515 N N Bacteroides coagulans 271 AB547639 clade_515N N Clostridiales bacterium 9403326 534 HM587324 clade_515 N NClostridiales bacterium ph2 539 JN837487 clade_515 N NPeptostreptococcus sp. 9succ1 1457 X90471 clade_515 N NPeptostreptococcus sp. oral clone AP24 1459 AB175072 clade_515 N NTissierella praeacuta 1924 NR_044860 clade_515 N N Helicobactercanadensis 994 ABQS01000108 clade_518 N N Peptostreptococcus anaerobius1455 AY326462 clade_520 N N Peptostreptococcus stomatis 1461ADGQ01000048 clade_520 N N Bilophila wadsworthia 367 ADCP01000166clade_521 N N Desulfovibrio vulgaris 761 NR_074897 clade_521 N NActinomyces nasicola 64 AJ508455 clade_523 N N Cellulosimicrobium funkei500 AY501364 clade_523 N N Lactococcus raffinolactis 1146 NR_044359clade_524 N N Bacteroidales genomosp. P1 258 AY341819 clade_529 N NBacteroidales genomosp. P2 oral clone MB1_G13 259 DQ003613 clade_529 N NBacteroidales genomosp. P3 oral clone MB1_G34 260 DQ003615 clade_529 N NBacteroidales genomosp. P4 oral clone MB2_G17 261 DQ003617 clade_529 N NBacteroidales genomosp. P5 oral clone MB2_P04 262 DQ003619 clade_529 N NBacteroidales genomosp. P6 oral clone MB3_C19 263 DQ003634 clade_529 N NBacteroidales genomosp. P8 oral clone MB4_G15 265 DQ003626 clade_529 N NBacteroidetes bacterium oral taxon D27 333 HM099638 clade_530 N NBacteroidetes bacterium oral taxon F31 334 HM099643 clade_530 N NBacteroidetes bacterium oral taxon F44 335 HM099649 clade_530 N NFlavobacterium sp. NF2_1 885 FJ195988 clade_530 N N Myroidesodoratimimus 1326 NR_042354 clade_530 N N Myroides sp. MY15 1327GU253339 clade_530 N N Chlamydiales bacterium NS16 507 JN606076clade_531 N N Chlamydophila pecorum 508 D88317 clade_531 N OPParachlamydia sp. UWE25 1423 BX908798 clade_531 N N Fusobacterium russii903 NR_044687 clade_532 N N Streptobacillus moniliformis 1784 NR_027615clade_532 N N Eubacteriaceae bacterium P4P_50 P4 833 AY207060 clade_533N N Abiotrophia defectiva 1 ACIN02000016 clade_534 N N Abiotrophia sp.oral clone P4PA_155 P1 3 AY207063 clade_534 N N Catonella genomosp. P1oral clone MB5_P12 496 DQ003629 clade_534 N N Catonella morbi 497ACIL02000016 clade_534 N N Catonella sp. oral clone FL037 498 AY349369clade_534 N N Eremococcus coleocola 818 AENN01000008 clade_534 N NFacklamia hominis 879 Y10772 clade_534 N N Granulicatella sp. M658_99_3962 AJ271861 clade_534 N N Campylobacter coli 459 AAFL01000004 clade_535N OP Campylobacter concisus 460 CP000792 clade_535 N OP Campylobacterfetus 462 ACLG01001177 clade_535 N OP Campylobacter jejuni 465 AL139074clade_535 N Category-B Campylobacter upsaliensis 473 AEPU01000040clade_535 N OP Atopobium minutum 183 HM007583 clade_539 N N Atopobiumparvulum 184 CP001721 clade_539 N N Atopobium rimae 185 ACFE01000007clade_539 N N Atopobium sp. BS2 186 HQ616367 clade_539 N N Atopobium sp.F0209 187 EU592966 clade_539 N N Atopobium sp. ICM42b10 188 HQ616393clade_539 N N Atopobium sp. ICM57 189 HQ616400 clade_539 N N Atopobiumvaginae 190 AEDQ01000024 clade_539 N N Coriobacteriaceae bacteriumBV3Ac1 677 JN809768 clade_539 N N Actinomyces naeslundii 63 X81062clade_54 N N Actinomyces oricola 67 NR_025559 clade_54 N N Actinomycesoris 69 BABV01000070 clade_54 N N Actinomyces sp. 7400942 70 EU484334clade_54 N N Actinomyces sp. ChDC B197 72 AF543275 clade_54 N NActinomyces sp. GEJ15 73 GU561313 clade_54 N N Actinomyces sp.M2231_94_1 79 AJ234063 clade_54 N N Actinomyces sp. oral clone GU067 83AY349362 clade_54 N N Actinomyces sp. oral clone IO077 85 AY349364clade_54 N N Actinomyces sp. oral clone IP073 86 AY349365 clade_54 N NActinomyces sp. oral clone JA063 88 AY349367 clade_54 N N Actinomycessp. oral taxon 170 89 AFBL01000010 clade_54 N N Actinomyces sp. oraltaxon 171 90 AECW01000034 clade_54 N N Actinomyces urogenitalis 95ACFH01000038 clade_54 N N Actinomyces viscosus 96 ACRE01000096 clade_54N N Orientia tsutsugamushi 1383 AP008981 clade_541 N OP Megamonasfuniformis 1198 AB300988 clade_542 N N Megamonas hypermegale 1199AJ420107 clade_542 N N Aeromicrobium marinum 102 NR_025681 clade_544 N NAeromicrobium sp. JC14 103 JF824798 clade_544 N N Luteococcus sanguinis1190 NR_025507 clade_544 N N Propionibacteriaceae bacterium NML 02_02651568 EF599122 clade_544 N N Rhodococcus corynebacterioides 1622 X80615clade_546 N N Rhodococcus erythropolis 1624 ACNO01000030 clade_546 N NRhodococcus fascians 1625 NR_037021 clade_546 N N Segniliparus rotundus1690 CP001958 clade_546 N N Segniliparus rugosus 1691 ACZI01000025clade_546 N N Exiguobacterium acetylicum 878 FJ970034 clade_547 N NMacrococcus caseolyticus 1194 NR_074941 clade_547 N N Streptomyces sp. 1AIP_2009 1890 FJ176782 clade_548 N N Streptomyces sp. SD 524 1892EU544234 clade_548 N N Streptomyces sp. SD 528 1893 EU544233 clade_548 NN Streptomyces thermoviolaceus 1895 NR_027616 clade_548 N N Borreliaafzelii 388 ABCU01000001 clade_549 N OP Borrelia crocidurae 390 DQ057990clade_549 N OP Borrelia duttonii 391 NC_011229 clade_549 N OP Borreliahermsii 393 AY597657 clade_549 N OP Borrelia hispanica 394 DQ057988clade_549 N OP Borrelia persica 395 HM161645 clade_549 N OP Borreliarecurrentis 396 AF107367 clade_549 N OP Borrelia spielmanii 398ABKB01000002 clade_549 N OP Borrelia turicatae 399 NC_008710 clade_549 NOP Borrelia valaisiana 400 ABCY01000002 clade_549 N OP Providenciaalcalifaciens 1586 ABXW01000071 clade_55 N N Providencia rettgeri 1587AM040492 clade_55 N N Providencia rustigianii 1588 AM040489 clade_55 N NProvidencia stuartii 1589 AF008581 clade_55 N N Treponema pallidum 1932CP001752 clade_550 N OP Treponema phagedenis 1934 AEFH01000172 clade_550N N Treponema sp. clone DDKL_4 1939 Y08894 clade_550 N N Acholeplasmalaidlawii 17 NR_074448 clade_551 N N Mycoplasma putrefaciens 1323 U26055clade_551 N N Mycoplasmataceae genomosp. P1 oral clone MB1_G23 1325DQ003614 clade_551 N N Spiroplasma insolitum 1750 NR_025705 clade_551 NN Collinsella intestinalis 660 ABXH02000037 clade_553 N N Collinsellastercoris 661 ABXJ01000150 clade_553 N N Collinsella tanakaei 662AB490807 clade_553 N N Caminicella sporogenes 458 NR_025485 clade_554 NN Acidaminococcus fermentans 21 CP001859 clade_556 N N Acidaminococcusintestini 22 CP003058 clade_556 N N Acidaminococcus sp. D21 23ACGB01000071 clade_556 N N Phascolarctobacterium faecium 1462 NR_026111clade_556 N N Phascolarctobacterium sp. YIT 12068 1463 AB490812clade_556 N N Phascolarctobacterium succinatutens 1464 AB490811clade_556 N N Acidithiobacillus ferrivorans 25 NR_074660 clade_557 N NXanthomonadaceae bacterium NML 03_0222 2015 EU313791 clade_557 N NCatabacter hongkongensis 494 AB671763 clade_558 N N Christensenellaminuta 512 AB490809 clade_558 N N Clostridiales bacterium oral cloneP4PA_66 P1 536 AY207065 clade_558 N N Clostridiales bacterium oral taxon093 537 GQ422712 clade_558 N N Heliobacterium modesticaldum 1000NR_074517 clade_560 N N Alistipes indistinctus 130 AB490804 clade_561 NN Bacteroidales bacterium ph8 257 JN837494 clade_561 N N CandidatesSulcia muelleri 475 CP002163 clade_561 N N Cytophaga xylanolytica 742FR733683 clade_561 N N Flavobacteriaceae genomosp. C1 884 AY278614clade_561 N N Gramella forsetii 958 NR_074707 clade_561 N NSphingobacterium faecium 1740 NR_025537 clade_562 N N Sphingobacteriummizutaii 1741 JF708889 clade_562 N N Sphingobacterium multivorum 1742NR_040953 clade_562 N N Sphingobacterium spiritivorum 1743 ACHA02000013clade_562 N N Jonquetella anthropi 1017 ACOO02000004 clade_563 N NPyramidobacter piscolens 1614 AY207056 clade_563 N N Synergistesgenomosp. C1 1904 AY278615 clade_563 N N Synergistes sp. RMA 14551 1905DQ412722 clade_563 N N Synergistetes bacterium ADV897 1906 GQ258968clade_563 N N Candidates Arthromitus sp. 474 NR_074460 clade_564 N NSFB_mouse_Yit Gracilibacter thermotolerans 957 NR_043559 clade_564 N NBrachyspira aalborgi 404 FM178386 clade_565 N N Brachyspira sp. HIS3 406FM178387 clade_565 N N Brachyspira sp. HIS4 407 FM178388 clade_565 N NBrachyspira sp. HIS5 408 FM178389 clade_565 N N Adlercreutziaequolifaciens 97 AB306661 clade_566 N N Coriobacteriaceae bacteriumJC110 678 CAEM01000062 clade_566 N N Coriobacteriaceae bacterium phI 679JN837493 clade_566 N N Cryptobacterium curtum 740 GQ422741 clade_566 N NEggerthella sinensis 779 AY321958 clade_566 N N Eggerthella sp.1_3_56FAA 780 ACWN01000099 clade_566 N N Eggerthella sp. HGA1 781AEXR01000021 clade_566 N N Eggerthella sp. YY7918 782 AP012211 clade_566N N Gordonibacter pamelaeae 680 AM886059 clade_566 N N Gordonibacterpamelaeae 956 FP929047 clade_566 N N Slackia equolifaciens 1732 EU377663clade_566 N N Slackia exigua 1733 ACUX01000029 clade_566 N N Slackiafaecicanis 1734 NR_042220 clade_566 N N Slackia heliotrinireducens 1735NR_074439 clade_566 N N Slackia isoflavoniconvertens 1736 AB566418clade_566 N N Slackia piriformis 1737 AB490806 clade_566 N N Slackia sp.NATTS 1738 AB505075 clade_566 N N Chlamydiales bacterium NS13 506JN606075 clade_567 N N Victivallaceae bacterium NML 080035 2003 FJ394915clade_567 N N Victivallis vadensis 2004 ABDE02000010 clade_567 N NStreptomyces griseus 1889 NR_074787 clade_573 N N Streptomyces sp. SD511 1891 EU544231 clade_573 N N Streptomyces sp. SD 534 1894 EU544232clade_573 N N Cloacibacillus evryensis 530 GQ258966 clade_575 N NDeferribacteres sp. oral clone JV001 743 AY349370 clade_575 N NDeferribacteres sp. oral clone JV023 745 AY349372 clade_575 N NSynergistetes bacterium LBVCM1157 1907 GQ258969 clade_575 N NSynergistetes bacterium oral taxon 362 1909 GU410752 clade_575 N NSynergistetes bacterium oral taxon D48 1910 GU430992 clade_575 N NPeptococcus sp. oral clone JM048 1439 AY349389 clade_576 N NHelicobacter winghamensis 999 ACDO01000013 clade_577 N N Wolinellasuccinogenes 2014 BX571657 clade_577 N N Olsenella genomosp. C1 1368AY278623 clade_578 N N Olsenella profusa 1369 FN178466 clade_578 N NOlsenella sp. F0004 1370 EU592964 clade_578 N N Olsenella sp. oral taxon809 1371 ACVE01000002 clade_578 N N Olsenella uli 1372 CP002106clade_578 N N Nocardiopsis dassonvillei 1356 CP002041 clade_579 N NPeptococcus niger 1438 NR_029221 clade_580 N N Peptococcus sp. oraltaxon 167 1440 GQ422727 clade_580 N N Akkermansia muciniphila 118CP001071 clade_583 N N Opitutus terrae 1373 NR_074978 clade_583 N NClostridiales bacterium oral taxon F32 538 HM099644 clade_584 N NLeptospira borgpetersenii 1161 NC_008508 clade_585 N OP Leptospirabroomii 1162 NR_043200 clade_585 N OP Leptospira interrogans 1163NC_005823 clade_585 N OP Methanobrevibacter gottschalkii 1213 NR_044789clade_587 N N Methanobrevibacter millerae 1214 NR_042785 clade_587 N NMethanobrevibacter oralis 1216 HE654003 clade_587 N N Methanobrevibacterthaueri 1219 NR_044787 clade_587 N N Methanobrevibacter smithii 1218ABYV02000002 clade_588 N N Deinococcus radiodurans 746 AE000513clade_589 N N Deinococcus sp. R_43890 747 FR682752 clade_589 N N Thermusaquaticus 1923 NR_025900 clade_589 N N Actinomyces sp. c109 81 AB167239clade_590 N N Syntrophomonadaceae genomosp. P1 1912 AY341821 clade_590 NN Anaerobaculum hydrogeniformans 141 ACJX02000009 clade_591 N NMicrocystis aeruginosa 1246 NC_010296 clade_592 N N Prochlorococcusmarinus 1567 CP000551 clade_592 N N Methanobrevibacter acididurans 1208NR_028779 clade_593 N N Methanobrevibacter arboriphilus 1209 NR_042783clade_593 N N Methanobrevibacter curvatus 1210 NR_044796 clade_593 N NMethanobrevibacter cuticularis 1211 NR_044776 clade_593 N NMethanobrevibacter filiformis 1212 NR_044801 clade_593 N NMethanobrevibacter woesei 1220 NR_044788 clade_593 N N Roseiflexuscastenholzii 1642 CP000804 clade_594 N N Methanobrevibacter olleyae 1215NR_043024 clade_595 N N Methanobrevibacter ruminantium 1217 NR_042784clade_595 N N Methanobrevibacter wolinii 1221 NR_044790 clade_595 N NMethanosphaera stadtmanae 1222 AY196684 clade_595 N N Chloroflexigenomosp. P1 511 AY331414 clade_596 N N Halorubrum lipolyticum 992AB477978 clade_597 N N Methanobacterium formicicum 1207 NR_025028clade_597 N N Acidilobus saccharovorans 24 AY350586 clade_598 N NHyperthermus butylicus 1006 CP000493 clade_598 N N Ignicoccus islandicus1011 X99562 clade_598 N N Metallosphaera sedula 1206 D26491 clade_598 NN Thermofilum pendens 1922 X14835 clade_598 N N Prevotellamelaninogenica 1506 CP002122 clade_6 N N Prevotella sp. ICM1 1520HQ616385 clade_6 N N Prevotella sp. oral clone FU048 1535 AY349393clade_6 N N Prevotella sp. oral clone GI030 1537 AY349395 clade_6 N NPrevotella sp. SEQ116 1526 JN867246 clade_6 N N Streptococcus anginosus1787 AECT01000011 clade_60 N N Streptococcus milleri 1812 X81023clade_60 N N Streptococcus sp. 16362 1829 JN590019 clade_60 N NStreptococcus sp. 69130 1832 X78825 clade_60 N N Streptococcus sp. AC151833 HQ616356 clade_60 N N Streptococcus sp. CM7 1839 HQ616373 clade_60N N Streptococcus sp. OBRC6 1847 HQ616352 clade_60 N N Burkholderiaambifaria 442 AAUZ01000009 clade_61 N OP Burkholderia cenocepacia 443AAHI01000060 clade_61 N OP Burkholderia cepacia 444 NR_041719 clade_61 NOP Burkholderia mallei 445 CP000547 clade_61 N Category-B Burkholderiamultivorans 446 NC_010086 clade_61 N OP Burkholderia oklahomensis 447DQ108388 clade_61 N OP Burkholderia pseudomallei 448 CP001408 clade_61 NCategory-B Burkholderia rhizoxinica 449 HQ005410 clade_61 N OPBurkholderia sp. 383 450 CP000151 clade_61 N OP Burkholderia xenovorans451 U86373 clade_61 N OP Prevotella buccae 1488 ACRB01000001 clade_62 NN Prevotella genomosp. P8 oral clone MB3_P13 1498 DQ003622 clade_62 N NPrevotella sp. oral clone FW035 1536 AY349394 clade_62 N N Prevotellabivia 1486 ADFO01000096 clade_63 N N Prevotella disiens 1494AEDO01000026 clade_64 N N Bacteroides faecis 276 GQ496624 clade_65 N NBacteroides fragilis 279 AP006841 clade_65 N N Bacteroides nordii 285NR_043017 clade_65 N N Bacteroides salyersiae 292 EU136690 clade_65 N NBacteroides sp. 1_1_14 293 ACRP01000155 clade_65 N N Bacteroides sp.1_1_6 295 ACIC01000215 clade_65 N N Bacteroides sp. 2_1_56FAA 298ACWI01000065 clade_65 N N Bacteroides sp. AR29 316 AF139525 clade_65 N NBacteroides sp. B2 317 EU722733 clade_65 N N Bacteroidesthetaiotaomicron 328 NR_074277 clade_65 N N Actinobacillus minor 45ACFT01000025 clade_69 N N Haemophilias parasuis 978 GU226366 clade_69 NN Vibrio cholerae 1996 AAUR01000095 clade_71 N Category-B Vibriofluvialis 1997 X76335 clade_71 N Category-B Vibrio furnissii 1998CP002377 clade_71 N Category-B Vibrio mimicus 1999 ADAF01000001 clade_71N Category-B Vibrio parahaemolyticus 2000 AAWQ01000116 clade_71 NCategory-B Vibrio sp. RC341 2001 ACZT01000024 clade_71 N Category-BVibrio vulnificus 2002 AE016796 clade_71 N Category-B Lactobacillusacidophilus 1067 CP000033 clade_72 N N Lactobacillus amylolyticus 1069ADNY01000006 clade_72 N N Lactobacillus amylovorus 1070 CP002338clade_72 N N Lactobacillus crispatus 1078 ACOG01000151 clade_72 N NLactobacillus delbrueckii 1080 CP002341 clade_72 N N Lactobacillushelveticus 1088 ACLM01000202 clade_72 N N Lactobacillus kalixensis 1094NR_029083 clade_72 N N Lactobacillus kefiranofaciens 1095 NR_042440clade_72 N N Lactobacillus leichmannii 1098 JX986966 clade_72 N NLactobacillus sp. 66c 1120 FR681900 clade_72 N N Lactobacillus sp. KLDS1.0701 1122 EU600905 clade_72 N N Lactobacillus sp. KLDS 1.0712 1130EU600916 clade_72 N N Lactobacillus sp. oral clone HT070 1136 AY349383clade_72 N N Lactobacillus ultunensis 1139 ACGU01000081 clade_72 N NPrevotella intermedia 1502 AF414829 clade_81 N N Prevotella nigrescens1511 AFPX01000069 clade_81 N N Prevotella pallens 1515 AFPY01000135clade_81 N N Prevotella sp. oral taxon 310 1551 GQ422737 clade_81 N NPrevotella genomosp. C1 1495 AY278624 clade_82 N N Prevotella sp. CM381519 HQ610181 clade_82 N N Prevotella sp. oral taxon 317 1552ACQH01000158 clade_82 N N Prevotella sp. SG12 1527 GU561343 clade_82 N NPrevotella denticola 1493 CP002589 clade_83 N N Prevotella genomosp. P7oral clone MB2_P31 1497 DQ003620 clade_83 N N Prevotella histicola 1501JN867315 clade_83 N N Prevotella multiformis 1508 AEWX01000054 clade_83N N Prevotella sp. JCM 6330 1522 AB547699 clade_83 N N Prevotella sp.oral clone GI059 1539 AY349397 clade_83 N N Prevotella sp. oral taxon782 1555 GQ422745 clade_83 N N Prevotella sp. oral taxon G71 1559GU432180 clade_83 N N Prevotella sp. SEQ065 1524 JN867234 clade_83 N NPrevotella veroralis 1565 ACVA01000027 clade_83 N N Bacteroidesacidifaciens 266 NR_028607 clade_85 N N Bacteroides cellulosilyticus 269ACCH01000108 clade_85 N N Bacteroides clarus 270 AFBM01000011 clade_85 NN Bacteroides eggerthii 275 ACWG01000065 clade_85 N N Bacteroidesoleiciplenus 286 AB547644 clade_85 N N Bacteroides pyogenes 290NR_041280 clade_85 N N Bacteroides sp. 315_5 300 FJ848547 clade_85 N NBacteroides sp. 31SF15 301 AJ583248 clade_85 N N Bacteroides sp. 31SF18302 AJ583249 clade_85 N N Bacteroides sp. 35AE31 303 AJ583244 clade_85 NN Bacteroides sp. 35AE37 304 AJ583245 clade_85 N N Bacteroides sp.35BE34 305 AJ583246 clade_85 N N Bacteroides sp. 35BE35 306 AJ583247clade_85 N N Bacteroides sp. WH2 324 AY895180 clade_85 N N Bacteroidessp. XB12B 325 AM230648 clade_85 N N Bacteroides stercoris 327ABFZ02000022 clade_85 N N Actinobacillus pleuropneumoniae 46 NR_074857clade_88 N N Actinobacillus ureae 48 AEVG01000167 clade_88 N NHaemophilus aegyptius 969 AFBC01000053 clade_88 N N Haemophilus ducreyi970 AE017143 clade_88 N OP Haemophilus haemolyticus 973 JN175335clade_88 N N Haemophilus influenzae 974 AADP01000001 clade_88 N OPHaemophilus parahaemolyticus 975 GU561425 clade_88 N N Haemophilusparainfluenzae 976 AEWU01000024 clade_88 N N Haemophilusparaphrophaemolyticus 977 M75076 clade_88 N N Haemophilus somnus 979NC_008309 clade_88 N N Haemophilus sp. 70334 980 HQ680854 clade_88 N NHaemophilus sp. HK445 981 FJ685624 clade_88 N N Haemophilus sp. oralclone ASCA07 982 AY923117 clade_88 N N Haemophilus sp. oral clone ASCG06983 AY923147 clade_88 N N Haemophilus sp. oral clone BJ021 984 AY005034clade_88 N N Haemophilus sp. oral clone BJ095 985 AY005033 clade_88 N NHaemophilus sp. oral taxon 851 987 AGRK01000004 clade_88 N N Haemophilussputorum 988 AFNK01000005 clade_88 N N Histophilus somni 1003 AF549387clade_88 N N Mannheimia haemolytica 1195 ACZX01000102 clade_88 N NPasteurella bettyae 1433 L06088 clade_88 N N Moellerella wisconsensis1253 JN175344 clade_89 N N Morganella morganii 1265 AJ301681 clade_89 NN Morganella sp. JB_T16 1266 AJ781005 clade_89 N N Proteus mirabilis1582 ACLE01000013 clade_89 N N Proteus penneri 1583 ABVP01000020clade_89 N N Proteus sp. HS7514 1584 DQ512963 clade_89 N N Proteusvulgaris 1585 AJ233425 clade_89 N N Oribacterium sinus 1374 ACKX1000142clade_90 N N Oribacterium sp. ACB1 1375 HM120210 clade_90 N NOribacterium sp. ACB7 1376 HM120211 clade_90 N N Oribacterium sp. CM121377 HQ616374 clade_90 N N Oribacterium sp. ICM51 1378 HQ616397 clade_90N N Oribacterium sp. OBRC12 1379 HQ616355 clade_90 N N Oribacterium sp.oral taxon 108 1382 AFIH01000001 clade_90 N N Actinobacillusactinomycetemcomitans 44 AY362885 clade_92 N N Actinobacillussuccinogenes 47 CP000746 clade_92 N N Aggregatibacteractinomycetemcomitans 112 CP001733 clade_92 N N Aggregatibacteraphrophilus 113 CP001607 clade_92 N N Aggregatibacter segnis 114AEPS01000017 clade_92 N N Averyella dalhousiensis 194 DQ481464 clade_92N N Bisgaard Taxon 368 AY683487 clade_92 N N Bisgaard Taxon 369 AY683489clade_92 N N Bisgaard Taxon 370 AY683491 clade_92 N N Bisgaard Taxon 371AY683492 clade_92 N N Buchnera aphidicola 440 NR_074609 clade_92 N NCedecea davisae 499 AF493976 clade_92 N N Citrobacter amalonaticus 517FR870441 clade_92 N N Citrobacter braakii 518 NR_028687 clade_92 N NCitrobacter farmeri 519 AF025371 clade_92 N N Citrobacter freundii 520NR_028894 clade_92 N N Citrobacter gillenii 521 AF025367 clade_92 N NCitrobacter koseri 522 NC_009792 clade_92 N N Citrobacter murliniae 523AF025369 clade_92 N N Citrobacter rodentium 524 NR_074903 clade_92 N NCitrobacter sedlakii 525 AF025364 clade_92 N N Citrobacter sp. 30_2 526ACDJ01000053 clade_92 N N Citrobacter sp. KMSI_3 527 GQ468398 clade_92 NN Citrobacter werkmanii 528 AF025373 clade_92 N N Citrobacter youngae529 ABWL02000011 clade_92 N N Cronobacter malonaticus 737 GU122174clade_92 N N Cronobacter sakazakii 738 NC_009778 clade_92 N NCronobacter turicensis 739 FN543093 clade_92 N N Enterobacter aerogenes786 AJ251468 clade_92 N N Enterobacter asburiae 787 NR_024640 clade_92 NN Enterobacter cancerogenus 788 Z96078 clade_92 N N Enterobacter cloacae789 FP929040 clade_92 N N Enterobacter cowanii 790 NR_025566 clade_92 NN Enterobacter hormaechei 791 AFHR01000079 clade_92 N N Enterobacter sp.247BMC 792 HQ122932 clade_92 N N Enterobacter sp. 638 793 NR_074777clade_92 N N Enterobacter sp. JC163 794 JN657217 clade_92 N NEnterobacter sp. SCSS 795 HM007811 clade_92 N N Enterobacter sp. TSE38796 HM156134 clade_92 N N Enterobacteriaceae bacterium 9_2_54FAA 797ADCU01000033 clade_92 N N Enterobacteriaceae bacterium CF01Ent_1 798AJ489826 clade_92 N N Enterobacteriaceae bacterium Smarlab 3302238 799AY538694 clade_92 N N Escherichia albertii 824 ABKX01000012 clade_92 N NEscherichia coli 825 NC_008563 clade_92 N Category-B Escherichiafergusonii 826 CU928158 clade_92 N N Escherichia hermannii 827 HQ407266clade_92 N N Escherichia sp. 1_1_43 828 ACID01000033 clade_92 N NEscherichia sp. 4_1_40B 829 ACDM02000056 clade_92 N N Escherichia sp. B4830 EU722735 clade_92 N N Escherichia vulneris 831 NR_041927 clade_92 NN Ewingella americana 877 JN175329 clade_92 N N Haemophilus genomosp. P2oral clone MB3_C24 971 DQ003621 clade_92 N N Haemophilus genomosp. P3oral clone MB3_C38 972 DQ003635 clade_92 N N Haemophilus sp. oral cloneJM053 986 AY349380 clade_92 N N Hafnia alvei 989 DQ412565 clade_92 N NKlebsiella oxytoca 1024 AY292871 clade_92 N OP Klebsiella pneumoniae1025 CP000647 clade_92 N OP Klebsiella sp. AS10 1026 HQ616362 clade_92 NN Klebsiella sp. Co9935 1027 DQ068764 clade_92 N N Klebsiella sp.enrichment culture clone SRC_DSD25 1036 HM195210 clade_92 N N Klebsiellasp. OBRC7 1028 HQ616353 clade_92 N N Klebsiella sp. SP_BA 1029 FJ999767clade_92 N N Klebsiella sp. SRC_DSD1 1033 GU797254 clade_92 N NKlebsiella sp. SRC_DSD11 1030 GU797263 clade_92 N N Klebsiella sp.SRC_DSD12 1031 GU797264 clade_92 N N Klebsiella sp. SRC_DSD15 1032GU797267 clade_92 N N Klebsiella sp. SRC_DSD2 1034 GU797253 clade_92 N NKlebsiella sp. SRC_DSD6 1035 GU797258 clade_92 N N Klebsiella variicola1037 CP001891 clade_92 N N Kluyvera ascorbata 1038 NR_028677 clade_92 NN Kluyvera cryocrescens 1039 NR_028803 clade_92 N N Leminorellagrimontii 1159 AJ233421 clade_92 N N Leminorella richardii 1160 HF558368clade_92 N N Pantoea agglomerans 1409 AY335552 clade_92 N N Pantoeaananatis 1410 CP001875 clade_92 N N Pantoea brenneri 1411 EU216735clade_92 N N Pantoea citrea 1412 EF688008 clade_92 N N Pantoea conspicua1413 EU216737 clade_92 N N Pantoea septica 1414 EU216734 clade_92 N NPasteurella dagmatis 1434 ACZR01000003 clade_92 N N Pasteurellamultocida 1435 NC_002663 clade_92 N N Plesiomonas shigelloides 1469X60418 clade_92 N N Raoultella ornithinolytica 1617 AB364958 clade_92 NN Raoultella planticola 1618 AF129443 clade_92 N N Raoultella terrigena1619 NR_037085 clade_92 N N Salmonella bongori 1683 NR_041699 clade_92 NCategory-B Salmonella enterica 1672 NC_011149 clade_92 N Category-BSalmonella enterica 1673 NC_011205 clade_92 N Category-B Salmonellaenterica 1674 DQ344532 clade_92 N Category-B Salmonella enterica 1675ABEH02000004 clade_92 N Category-B Salmonella enterica 1676 ABAK02000001clade_92 N Category-B Salmonella enterica 1677 NC_011080 clade_92 NCategory-B Salmonella enterica 1678 EU118094 clade_92 N Category-BSalmonella enterica 1679 NC_011094 clade_92 N Category-B Salmonellaenterica 1680 AE014613 clade_92 N Category-B Salmonella enterica 1682ABFH02000001 clade_92 N Category-B Salmonella enterica 1684 ABEM01000001clade_92 N Category-B Salmonella enterica 1685 ABAM02000001 clade_92 NCategory-B Salmonella typhimurium 1681 DQ344533 clade_92 N Category-BSalmonella typhimurium 1686 AF170176 clade_92 N Category-B Serratiafonticola 1718 NR_025339 clade_92 N N Serratia liquefaciens 1719NR_042062 clade_92 N N Serratia marcescens 1720 GU826157 clade_92 N NSerratia odorifera 1721 ADBY01000001 clade_92 N N Serratiaproteamaculans 1722 AAUN01000015 clade_92 N N Shigella boydii 1724AAKA01000007 clade_92 N Category-B Shigella dysenteriae 1725 NC_007606clade_92 N Category-B Shigella flexneri 1726 AE005674 clade_92 NCategory-B Shigella sonnei 1727 NC_007384 clade_92 N Category-BTatumella ptyseos 1916 NR_025342 clade_92 N N Trabulsiella guamensis1925 AYS73830 clade_92 N N Yersinia aldovae 2019 AJ871363 clade_92 N OPYersinia aleksiciae 2020 AJ627597 clade_92 N OP Yersinia bercovieri 2021AF366377 clade_92 N OP Yersinia enterocolitica 2022 FR729477 clade_92 NCategory-B Yersinia frederiksenii 2023 AF366379 clade_92 N OP Yersiniaintermedia 2024 AF366380 clade_92 N OP Yersinia kristensenii 2025ACCA01000078 clade_92 N OP Yersinia mollaretii 2026 NR_027546 clade_92 NOP Yersinia pestis 2027 AE013632 clade_92 N Category-A Yersiniapseudotuberculosis 2028 NC_009708 clade_92 N OP Yersinia rohdei 2029ACCD01000071 clade_92 N OP Yokenella regensburgei 2030 AB273739 clade_92N N Conchiformibius kuhniae 669 NR_041821 clade_94 N N Morococcuscerebrosus 1267 JN175352 clade_94 N N Neisseria bacilliformis 1328AFAY01000058 clade_94 N N Neisseria cinerea 1329 ACDY01000037 clade_94 NN Neisseria flavescens 1331 ACQV01000025 clade_94 N N Neisseriagonorrhoeae 1333 CP002440 clade_94 N OP Neisseria lactamica 1334ACEQ01000095 clade_94 N N Neisseria macacae 1335 AFQE01000146 clade_94 NN Neisseria meningitidis 1336 NC_003112 clade_94 N OP Neisseria mucosa1337 ACDX01000110 clade_94 N N Neisseria pharyngis 1338 AJ239281clade_94 N N Neisseria polysaccharea 1339 ADBE01000137 clade_94 N NNeisseria sicca 1340 ACKO02000016 clade_94 N N Neisseria sp. KEM232 1341GQ203291 clade_94 N N Neisseria sp. oral clone AP132 1344 AY005027clade_94 N N Neisseria sp. oral strain B33KA 1346 AY005028 clade_94 N NNeisseria sp. oral taxon 014 1347 ADEA01000039 clade_94 N N Neisseriasp. TM10_1 1343 DQ279352 clade_94 N N Neisseria subflava 1348ACEO01000067 clade_94 N N Okadaella gastrococcus 1365 HQ699465 clade_98N N Streptococcus agalactiae 1785 AAJO01000130 clade_98 N NStreptococcus alactolyticus 1786 NR_041781 clade_98 N N Streptococcusaustralis 1788 AEQR01000024 clade_98 N N Streptococcus bovis 1789AEEL01000030 clade_98 N N Streptococcus canis 1790 AJ413203 clade_98 N NStreptococcus constellatus 1791 AY277942 clade_98 N N Streptococcuscristatus 1792 AEVC01000028 clade_98 N N Streptococcus dysgalactiae 1794AP010935 clade_98 N N Streptococcus equi 1795 CP001129 clade_98 N NStreptococcus equinus 1796 AEVB01000043 clade_98 N N Streptococcusgallolyticus 1797 FR824043 clade_98 N N Streptococcus genomosp. C1 1798AY278629 clade_98 N N Streptococcus genomosp. C2 1799 AY278630 clade_98N N Streptococcus genomosp. C3 1800 AY278631 clade_98 N N Streptococcusgenomosp. C4 1801 AY278632 clade_98 N N Streptococcus genomosp. C5 1802AY278633 clade_98 N N Streptococcus genomosp. C6 1803 AY278634 clade_98N N Streptococcus genomosp. C7 1804 AY278635 clade_98 N N Streptococcusgenomosp. C8 1805 AY278609 clade_98 N N Streptococcus gordonii 1806NC_009785 clade_98 N N Streptococcus infantarius 1807 ABJK02000017clade_98 N N Streptococcus infantis 1808 AFNN01000024 clade_98 N NStreptococcus intermedius 1809 NR_028736 clade_98 N N Streptococcuslutetiensis 1810 NR_037096 clade_98 N N Streptococcus massiliensis 1811AY769997 clade_98 N N Streptococcus mitis 1813 AM157420 clade_98 N NStreptococcus oligofermentans 1815 AY099095 clade_98 N N Streptococcusoralis 1816 ADMV01000001 clade_98 N N Streptococcus parasanguinis 1817AEKM01000012 clade_98 N N Streptococcus pasteurianus 1818 AP012054clade_98 N N Streptococcus peroris 1819 AEVF01000016 clade_98 N NStreptococcus pneumoniae 1820 AE008537 clade_98 N N Streptococcusporcinus 1821 EF121439 clade_98 N N Streptococcus pseudopneumoniae 1822FJ827123 clade_98 N N Streptococcus pseudoporcinus 1823 AENS01000003clade_98 N N Streptococcus pyogenes 1824 AE006496 clade_98 N OPStreptococcus ratti 1825 X58304 clade_98 N N Streptococcus sanguinis1827 NR_074974 clade_98 N N Streptococcus sinensis 1828 AF432857clade_98 N N Streptococcus sp. 2_1_36FAA 1831 ACOI01000028 clade_98 N NStreptococcus sp. 2285_97 1830 AJ131965 clade_98 N N Streptococcus sp.ACS2 1834 HQ616360 clade_98 N N Streptococcus sp. AS20 1835 HQ616366clade_98 N N Streptococcus sp. BS35a 1836 HQ616369 clade_98 N NStreptococcus sp. C150 1837 ACRI01000045 clade_98 N N Streptococcus sp.CM6 1838 HQ616372 clade_98 N N Streptococcus sp. ICM10 1840 HQ616389clade_98 N N Streptococcus sp. ICM12 1841 HQ616390 clade_98 N NStreptococcus sp. ICM2 1842 HQ616386 clade_98 N N Streptococcus sp. ICM41844 HQ616387 clade_98 N N Streptococcus sp. ICM45 1843 HQ616394clade_98 N N Streptococcus sp. M143 1845 ACRK01000025 clade_98 N NStreptococcus sp. M334 1846 ACRL01000052 clade_98 N N Streptococcus sp.oral clone ASB02 1849 AY923121 clade_98 N N Streptococcus sp. oral cloneASCA03 1850 DQ272504 clade_98 N N Streptococcus sp. oral clone ASCA041851 AY923116 clade_98 N N Streptococcus sp. oral clone ASCA09 1852AY923119 clade_98 N N Streptococcus sp. oral clone ASCB04 1853 AY923123clade_98 N N Streptococcus sp. oral clone ASCB06 1854 AY923124 clade_98N N Streptococcus sp. oral clone ASCC04 1855 AY923127 clade_98 N NStreptococcus sp. oral clone ASCC05 1856 AY923128 clade_98 N NStreptococcus sp. oral clone ASCC12 1857 DQ272507 clade_98 N NStreptococcus sp. oral clone ASCD01 1858 AY923129 clade_98 N NStreptococcus sp. oral clone ASCD09 1859 AY923130 clade_98 N NStreptococcus sp. oral clone ASCD10 1860 DQ272509 clade_98 N NStreptococcus sp. oral clone ASCE03 1861 AY923134 clade_98 N NStreptococcus sp. oral clone ASCE04 1862 AY953253 clade_98 N NStreptococcus sp. oral clone ASCE05 1863 DQ272510 clade_98 N NStreptococcus sp. oral clone ASCE06 1864 AY923135 clade_98 N NStreptococcus sp. oral clone ASCE09 1865 AY923136 clade_98 N NStreptococcus sp. oral clone ASCE10 1866 AY923137 clade_98 N NStreptococcus sp. oral clone ASCE12 1867 AY923138 clade_98 N NStreptococcus sp. oral clone ASCF05 1868 AY923140 clade_98 N NStreptococcus sp. oral clone ASCF07 1869 AY953255 clade_98 N NStreptococcus sp. oral clone ASCF09 1870 AY923142 clade_98 N NStreptococcus sp. oral clone ASCG04 1871 AY923145 clade_98 N NStreptococcus sp. oral clone BW009 1872 AY005042 clade_98 N NStreptococcus sp. oral clone CH016 1873 AY005044 clade_98 N NStreptococcus sp. oral clone GK051 1874 AY349413 clade_98 N NStreptococcus sp. oral clone GM006 1875 AY349414 clade_98 N NStreptococcus sp. oral clone P2PA_41 P2 1876 AY207051 clade_98 N NStreptococcus sp. oral clone P4PA_30 P4 1877 AY207064 clade_98 N NStreptococcus sp. oral taxon 071 1878 AEEP01000019 clade_98 N NStreptococcus sp. oral taxon G59 1879 GU432132 clade_98 N NStreptococcus sp. oral taxon G62 1880 GU432146 clade_98 N NStreptococcus sp. oral taxon G63 1881 GU432150 clade_98 N NStreptococcus suis 1882 FM252032 clade_98 N N Streptococcus thermophilus1883 CP000419 clade_98 N N Streptococcus salivarius 1826 AGBV01000001clade_98 N N Streptococcus uberis 1884 HQ391900 clade_98 N NStreptococcus urinalis 1885 DQ303194 clade_98 N N Streptococcusvestibularis 1886 AEKO01000008 clade_98 N N Streptococcus viridans 1887AF076036 clade_98 N N Synergistetes bacterium oral clone 03 5 D05 1908GU227192 clade_98 N N

TABLE 1A Exemplary Immunomodulatory Bacterial Species Alkaliphilusmetalliredigens Ammonifex degensii Anaerofustis stercorihominisAnaerostipes caccae Anaerotruncus colihominis Bacillus amyloliquefaciensBacillus anthracis Bacillus cellulosilyticus Bacillus cereus Bacillusclausii Bacillus coagulans Bacillus cytotoxicus Bacillus haloduransBacillus licheniformis Bacillus pumilus Bacillus subtilis Bacillusthuringiensis Bacillus weihenstephanensis Blautia (Ruminococcus)hansenii Blautia (Ruminococcus) obeum Brevibacillus brevis Bryantellaformatexigens Caldicellulosiruptor saccharolyticus CandidatusDesulforudis audaxviator Carboxydibrachium pacificum Carboxydothermushydrogenoformans Clostridium acetobutylicum Clostridium asparagiformeClostridium bartlettii Clostridium beijerinckii Clostridium bolteaeClostridium botulinum A str. ATCC 19397 Clostridium botulinum B str.Eklund 17B Clostridium butyricum pathogenic E4 str. BoNT BL5262Clostridium carboxidivorans Clostridium cellulolyticum Clostridiumcellulovorans Clostridium difficile Clostridium (Hungatella) hathewayiClostridium hylemonae Clostridium kluyveri Clostridium leptumClostridium methylpentosum Clostridium (Tyzzerella) nexile Clostridiumnovyi NT Clostridium papyrosolvens Clostridium perfringens Clostridiumphytofermentans ISDg Clostridium scindens Clostridium sp. 7_2_43FAAClostridium sporogenes Clostridium tetani Clostridium thermocellumCoprococcus comes Desulfotomaculum reducens Dorea longicatenaEubacterium eligens Eubacterium hallii Eubacterium rectale Eubacteriumventriosum Faecalibacterium prausnitzii Geobacillus kaustophilusGeobacillus sp. G11MC16 Geobacillus thermodenitrificans Heliobacteriummodesticaldum Lysinibacillus sphaericus Oceanobacillus iheyensisPaenibacillus sp. JDR-2 Pelotomaculum thermopropionicum Roseburiaintestinalis Ruminococcus bromii Ruminococcus gnavus Ruminococcustorques Subdoligranulum variabile Symbiobacterium thermophilumThermoanaerobacter italicus Thermoanaerobacter tengcongensisThermoanaerobacterium thermosaccharolyticum Thermosinus carboxydivorans

TABLE 1B Exemplary Bacteria Useful in the Present InventionAcidaminococcus intestini Acinetobacter baumannii Acinetobacter lwoffiiAkkermansia muciniphila Alistipes putredinis Alistipes shahiiAnaerostipes hadrus Anaerotruncus colihominis Bacteroides caccaeBacteroides cellulosilyticus Bacteroides dorei Bacteroides eggerthiiBacteroides finegoldii Bacteroides fragilis Bacteroides massiliensisBacteroides ovatus Bacteroides salanitronis Bacteroides salyersiaeBacteroides sp. 1_1_6 Bacteroides sp. 3_1_23 Bacteroides sp. D20Bacteroides thetaiotaomicron Bacteroides uniformis Bacteroides vulgatusBifidobacterium adolescentis Bifidobacterium bifidum Bifidobacteriumbreve Bifidobacterium faecale Bifidobacterium kashiwanohenseBifidobacterium longum subsp. longum Bifidobacterium pseudocatenulatumBifidobacterium stercoris Blautia (Ruminococcus) coccoides Blautiafaecis Blautia glucerasea Blautia (Ruminococcus) hansenii Blautiahydrogenotrophica (Ruminococcus hydrogenotrophicus) Blautia(Ruminococcus) luti Blautia (Ruminococcus) obeum Blautia producta(Ruminococcus productus) Blautia (Ruminococcus) schinkii Blautiastercoris Blautia uncultured bacterium clone BKLE_a03_2 (GenBank:EU469501.1) Blautia uncultured bacterium clone SJTU_B_14_30 (GenBank:EF402926.1) Blautia uncultured bacterium clone SJTU_C_14_16 (GenBank:EF404657.1) Blautia uncultured bacterium clone S1-5 (GenBank:GQ898099.1) Blautia uncultured PAC000178_s(www.ezbiocloud.net/eztaxon/hierarchy?m=browse&k=PAC000178&d=2) Blautiawexlerae Candidatus Arthromitus sp. SFB-mouse-Yit Catenibacteriummitsuokai Clostridiaceae bacterium (Dielma fastidiosa) JC13Clostridiales bacterium 1_7_47FAA Clostridium asparagiforme Clostridiumbolteae Clostridium clostridioforme Clostridium glycyrrhizinilyticumClostridium (Hungatella) hathewayi Clostridium histolyticum Clostridiumindolis Clostridium leptum Clostridium (Tyzzerella) nexile Clostridiumperfringens Clostridium (Erysipelatoclostridium) ramosum Clostridiumscindens Clostridium sp. 14774 Clostridium sp. 7_3_54FAA Clostridium sp.HGF2 Clostridium symbiosum Collinsella aerofaciens Collinsellaintestinalis Coprobacillus sp. D7 Coprococcus catus Coprococcus comesDorea formicigenerans Dorea longicatena Enterococcus faecalisEnterococcus faecium Erysipelotrichaceae bacterium 3_1_53 Escherichiacoli Escherichia coli S88 Eubacterium eligens Eubacterium fissicatenaEubacterium ramulus Eubacterium rectale Faecalibacterium prausnitziiFlavonifractor plautii Fusobacterium mortiferum Fusobacterium nucleatumHoldemania filiformis Hydrogenoanaerobacterium saccharovorans Klebsiellaoxytoca Lachnospiraceae bacterium 3_1_57FAA_CT1 Lachnospiraceaebacterium 7_1_58FAA Lachnospiraceae bacterium 5_1_57FAA Lactobacilluscasei Lactobacillus rhamnosus Lactobacillus ruminis Lactococcus caseiOdoribacter splanchnicus Oscillibacter valericigenes Parabacteroidesgordonii Parabacteroides johnsonii Parabacteroides merdae Pediococcusacidilactici Peptostreptococcus asaccharolyticus Propionibacteriumgranulosum Roseburia intestinalis Roseburia inulinivorans Ruminococcusfaecis Ruminococcus gnavus Ruminococcus sp. ID8 Ruminococcus torquesSlackia piriformis Staphylococcus epidermidis Staphylococcussaprophyticus Streptococcus cristatus Streptococcus dysgalactiae subsp.equisimilis Streptococcus infantis Streptococcus oralis Streptococcussanguinis Streptococcus viridans Streptococcus thermophilus Veillonelladispar

TABLE 1C Exemplary Bacteria Useful in the Present InventionAnaerotruncus colihominis strain 13 Blautia producta strain 6Clostridium bolteae strain 7 Clostridiaceae bacterium JC 13 strain 8Clostridiales bacterium 1_7_47FAA strain 28 Clostridium sp. 7_3_54FAAstrain 16 Clostridium asparagiforme strain 15 Clostridiumclostridioforme Clostridium (Hungatella) hatewayi strain 4 Clostridiumindolis strain 9 Clostridium (Erysipelatoclostridium) ramosum strain 18Clostridium scindens strain 26 Clostridium sp. 14774 strain 1Eubacterium fissicatena strain 21 Hydrogenoanaerobacteriumsaccharovorans Lachinospiraceae bacterium 3_1_57FAA strain 27Lachinospiraceae bacterium 3_1_57FAA strain 29 Lachinospiraceaebacterium 7_1_58FAA strain 3 Oscillibacter valericigens Ruminococcus sp.ID8 strain 14

TABLE 1D Exemplary Bacteria Useful in the Present Invention Bacteroidescaccae Bacteroides eggerthii Bacteroides ovatus Bacteroides sp. 1_1_6Bacteroides sp. 3_1_23 Bacteroides sp. D20 Bacteroides vulgatusBifidobacterium adolescentis Bifidobacterium pseudocatenulatum Blautia(Ruminococcus) obeum Blautia producta (Ruminococcus productus) Blautia(Ruminococcus) schinkii Clostridium (Hungatella) hathewayi Clostridium(Tyzzerella) nexile Clostridium sp. HGF2 Clostridium symbiosumCollinsella aerofaciens Coprobacillus sp. D7 Coprococcus catusCoprococcus comes Dorea formicigenerans Dorea longicatena Enterococcusfaecalis Erysipelotrichaceae bacterium 3_1_53 Escherichia coliEscherichia coli S88 Eubacterium eligens Eubacterium rectaleFaecalibacterium prausnitzii Lachnospiraceae bacterium 5_1_57FAAOdoribacter splanchnicus Parabacteroides merdae Roseburia intestinalisRuminococcus torques Streptococcus thermophilus

TABLE 1E Exemplary Bacteria Useful in the Present Invention Akkermansiamuciniphila Enterococcus faecalis Klebsiella oxytoca Lactobacillusrhamnosus Staphylococcus epidermidis Streptococcus viridans Veillonelladispar

TABLE 1F Exemplary Bacteria Useful in the Present InventionAcinetobacter baumannii Acinetobacter lwoffii Akkermansia muciniphilaAlistipes shahii Anaerotruncus colihominis Bacteroides caccaeBacteroides dorei Bacteroides eggerthii Bacteroides finegoldiiBacteroides fragilis Bacteroides massiliensis Bacteroides ovatusBacteroides salanitronis Bacteroides sp. 1_1_6 Bacteroides sp. 3_1_23Bacteroides sp. D20 Bacteroides thetaiotaomicron Bacteroides uniformisBacteroides vulgatus Bifidobacterium adolescentis Bifidobacterium breveBifidobacterium pseudocatenulatum Blautia (Ruminococcus) coccoidesBlautia faecis Blautia glucerasea Blautia (Ruminococcus) hanseniiBalutia hydrogenotrophica (Ruminococcus hydrogentrophicus) Blautia(Ruminococcus) luti Blautia (Ruminococcus) obeum Blautia producta(Ruminococcus productus) Blautia (Ruminococcus) schinkii Blautiastercoris Blautia wexlerae Candidatus Arthromitus sp. SFB-mouse-YitClostridiaceae bacterium (Dielma fastidiosa) JC13 Clostridialesbacterium 1_7_47FAA Clostridium asparagiforme Clostridium bolteaeClostridium clostridioforme Clostridium (Hungatella) hathewayiClostridium histolyticum Clostridium indolis Clostridium leptumClostridium (Tyzzerella) nexile Clostridium perfringens Clostridium(Erysipelatoclostridium) ramosum Clostridium scindens Clostridium sp.14774 Clostridium sp. 7_3_54FAA Clostridium sp. HGF2 Clostridiumsymbiosum Collinsella aerofaciens Coprobacillus sp. D7 Coprococcus catusCoprococcus comes Dorea formicigenerans Dorea longicatena Enterococcusfaecium Erysipelotrichaceae bacterium 3_1_53 Escherichia coliEscherichia coli S88 Eubacterium eligens Eubacterium fissicatenaEubacterium rectale Faecalibacterium prausnitzii Fusobacteriummortiferum Fusobacterium nucleatum Hydrogenoeneaerobacteriumsaccharovorans Lachnospiraceae bacterium 3_1_57FAA_CT1 Lachnospiraceaebacterium 7_1_58FAA Lachnospiraceae bacterium 5_1_57FAA Lactobacilluscasei Lactococcus casei Odoribacter splanchnicus Oscillibactervalericigenes Parabacteroides johnsonii Parabacteroides merdaePediococcus acidilactici Peptostreptococcus asaccharolyticusPropionibacterium granulosum Roseburia intestinalis Ruminococcus gnavusRuminococcus sp. ID8 Ruminococcus torques Staphylococcus saprophyticusStreptococcus thermophilus

TABLE 2A Species identified as germinable and sporulatable by colonypicking GAM + Sweet B + Sweet OTU BBA FOS/inulin M2GSC FOS/Inulin GAMTotal Blautia producta 1 1 Clostridium bartlettii 4 1 5 Clostridiumbolteae 2 5 1 8 Clostridium botulinum 5 5 Clostridium butyricum 37 43 81 33 122 Clostridium celatum 4 1 5 Clostridium clostridioforme 1 1 2Clostridium disporicum 26 26 22 33 50 157 Clostridium glycolicum 4 9 1427 Clostridium mayombei 2 2 4 Clostridium paraputrificum 8 8 33 16 6 71Clostridium sordellii 14 14 Clostridium sp. 7_2_43FAA 1 1 Clostridiumsymbiosum 3 3 Clostridium tertium 1 1 2 (blank) 2 31 33 Totals 92 92 9292 92 460

TABLE 2B Species identified as germinable by 16S colony pick approachClostridium paraputrificum Clostridium disporicum Clostridium glucolicumClostridium bartlettii Clostridium butyricum Ruminococcus bromiiLachnospiraceae bacterium 2_1_58FAA Eubacterium hadrum Turicibactersanguinis Lachnospiraceae bacterium oral taxon F15 Clostridiumperfringens Clostridium bifermentans Roseburia sp 11SE37 Clostridiumquinii Ruminococcus lactaris Clostridium botulinum Clostridiumtyrobutyricum Blautia hansenii Clostridium kluyveri Clostridium sp JC122Clostridium hylemonae Clostridium celatum Clostridium straminisolvensClostridium orbischindens Roseburia cecicola Eubacterium tenueClostridium sp. 7_2_43FAA Lachnospiraceae bacterium 4_1_37FAAEubacterium rectale Clostridium viride Ruminococcus sp. K_1 Clostridiumsymbiosum Ruminococcus torques Clostridium algidicarnis

TABLE 2C Species identified as sporulatable by 16S NGS approachClostridium paraputrificum Clostridium bartlettii Lachnospiraceaebacterium 2_1_58FAA Clostridium disporicum Ruminococcus bromiiEubacterium hadrum Clostridium butyricum Roseburia sp. 11SE37Clostridium perfringens Clostridium glycolicum Clostridium hylemonaeClostridium orbiscindens Ruminococcus lactaris Clostridium symbiosumLachnospiraceae bacterium oral taxon F15 Blautia hansenii Turicibactersanguinis Clostridium straminisolvens Clostridium botulinumLachnospiraceae bacterium 4_1_37FAA Roseburia cecicola Ruminococcus sp.K_1 Clostridium bifermentans Eubacterium rectale Clostridium quiniiClostridium viride Clostridium kluyveri Clostridium tyrobutyricumOscillibacter sp. G2 Clostridium sp. JC122 Lachnospiraceae bacterium3_1_57FAA Clostridium aldenense Ruminococcus torques Clostridium sp.7_2_43FAA Clostridium celatum Eubacterium sp. WAL_14571 Eubacteriumtenue Lachnospiraceae bacterium 5_1_57FAA Clostridium clostridioformeClostridium sp. YIT_12070 Blautia sp. M25 Anaerostipes caccae Roseburiainulinivorans Clostridium sp. D5 Clostridium aparagiforme Coprobacillussp. D7 Clostridium sp. HGF2 Clostridium citroniae Clostridium difficileOscillibacter valericigenes Clostridium algidicarnis

TABLE 3 Anaerobic bacterial species tested for carbon source usage(Biolog plates) Species purchased: Species Freshly Isolated: R. gnavus(EPV1) Blautia luti B1nIX (EPV114) E. rectale (EPV2) Blautia luti ELU(EPV54) B. luti (EPV3) Ruminococcus gnavus (EPV102) B. wexlerae (EPV5)Blautia faecis (EPV78) C. leptum (EPV6) Ruminococcus torques (EPV76) B.faecis (EPV15) Blautia wexlerae SJTU1416 (EPV52) B. obeum (EPV20)Blautia WAL14507 (EPV64) B. producta (EPV21) Uncultured bacteriumSJTU1416 (EPV51) B. coccoides (EPV22) Uncultured bacterium GQ8980099(EPV47) B. hydrogenotrophica (EPV23) Eubacterium rectale (EPV35) B.hansenii (EPV24)

TABLE 4 Exemplary Prebiotics/Carbon Sources Chemical MoA L-ArabinoseC-Source, carbohydrate N-Acetyl-D-Glucosamine C-Source, carbohydrateD-Saccharic acid C-Source, carboxylic acid Succinic acid C-Source,carboxylic acid D-Galactose C-Source, carbohydrate L-Aspartic acidC-Source, amino acid L-Proline C-Source, amino acid D-Alanine C-Source,amino acid D-Trehalose C-Source, carbohydrate D-Mannose C-Source,carbohydrate Dulcitol C-Source, carbohydrate D-Serine C-Source, aminoacid D-Sorbitol C-Source, carbohydrate Glycerol C-Source, carbohydrateL-Fucose C-Source, carbohydrate D-Glucuronic acid C-Source, carboxylicacid D-Gluconic acid C-Source, carboxylic acid DL-a-Glycerol PhosphateC-Source, carbohydrate D-Xylose C-Source, carbohydrate L-Lactic acidC-Source, carboxylic acid Formic acid C-Source, carboxylic acidD-Mannitol C-Source, carbohydrate L-Glutamic acid C-Source, amino acidD-Glucose-6-Phosphate C-Source, carbohydrate D-Galactonic acid-g-LactoneC-Source, carboxylic acid DL-Malic acid C-Source, carboxylic acidD-Ribose C-Source, carbohydrate Tween 20 C-Source, fatty acid L-RhamnoseC-Source, carbohydrate D-Fructose C-Source, carbohydrate Acetic acidC-Source, carboxylic acid a-D-Glucose C-Source, carbohydrate MaltoseC-Source, carbohydrate D-Melibiose C-Source, carbohydrate ThymidineC-Source, carbohydrate L-Asparagine C-Source, amino acid D-Aspartic acidC-Source, amino acid D-Glucosamine acid C-Source, carboxylic acid1,2-Propanediol C-Source, alcohol Tween 40 C-Source, fatty acida-Ketoglutaric acid C-Source, carboxylic acid a-Ketobutyric acidC-Source, carboxylic acid a-methyl-D-Galactoside C-Source, carbohydratea-D-Lactose C-Source, carbohydrate Lactulose C-Source, carbohydrateSucrose C-Source, carbohydrate Uridine C-Source, carbohydrateL-Glutamine C-Source, amino acid m-Tartaric acid C-Source, carboxylicacid D-Glucose-1-Phosphate C-Source, carbohydrate D-Fructose-6-PhosphateC-Source, carbohydrate Tween 80 C-Source, fatty acid a-Hydroxyglutaricacid-g-Lactone C-Source, carboxylic acid a-Hydroxybutyric acid C-Source,carboxylic acid b-Methyl-D-GLucoside C-Source, carbohydrate AdonitolC-Source, carbohydrate Maltotriose C-Source, carbohydrate2′-Deoxyadenosine C-Source, carbohydrate Adenosine C-Source,carbohydrate Gly-Asp C-Source, amino acid Citric acid C-Source,carboxylic acid m-Inositol C-Source, carbohydrate D-Threonine C-Source,amino acid Fumaric acid C-Source, carboxylic acid Bromosuccinic acidC-Source, carboxylic acid Propionic acid C-Source, carboxylic acid Mucicacid C-Source, carboxylic acid Glycolic acid C-Source, carboxylic acidGlyoxylic acid C-Source, carboxylic acid D-Celloniose C-Source,carbohydrate Inosine C-Source, carbohydrate Gly-Glu C-Source, amino acidTricarballylic acid C-Source, carboxylic acid L-Serine C-Source, aminoacid L-Threonine C-Source, amino acid L-Alanine C-Source, amino acidAla-Gly C-Source, amino acid Acetoacetic acid C-Source, carboxylic acidN-Acetyl-D-Mannosamine C-Source, carbohydrate Mono-MethylsuccinateC-Source, carboxylic acid Methylpyruvate C-Source, ester D-Malic acidC-Source, carboxylic acid L-Malic acid C-Source, carboxylic acid Gly-ProC-Source, amino acid p-Hydroxyphenyl Acetic acid C-Source, carboxylicacid m-Hydroxyphenyl Acetic acid C-Source, carboxylic acid TyramineC-Source, amine D-Psicose C-Source, carbohydrate L-Lyxose C-Source,carbohydrate Glucuronamide C-Source, amide Pyruvic acid C-Source,carboxylic acid L-Galactonic acid-g-Lactone C-Source, carboxylic acidD-Galacturonic acid C-Source, carboxylic acid Phenylethylamine C-Source,amine 2-Aminoethanol C-Source, alcohol Negative Control C-Source,negative control Chondroitin Sulfate C C-Source, polymer a-CyclodextrinC-Source, polymer b-Cyclodextrin C-Source, polymer g-CyclohextrinC-Source, polymer Dextrin C-Source, polymer Gelatin C-Source, polymerGlycogen C-Source, polymer Inulin C-Source, polymer Laminarin C-Source,polymer Mannan C-Source, polymer Pectin C-Source, polymerN-Acetyl-D-Galactosamine C-Source, carbohydrate N-Acetyl-Neuraminic acidC-Source, carboxylic acid b-D-Allose C-Source, carbohydrate AmygdalinC-Source, carbohydrate D-Arabinose C-Source, carbohydrate D-ArabitolC-Source, carbohydrate L-Arabitol C-Source, carbohydrate ArbutinC-Source, carbohydrate 2-Deoxy-D-Ribose C-Source, carbohydratei-Erythritol C-Source, carbohydrate D-Fucose C-Source, carbohydrate3-O-b-D-Galactopyranosyl-D-Arabinose C-Source, carbohydrate GentiobioseC-Source, carbohydrate L-Glucose C-Source, carbohydrate D-LactitolC-Source, carbohydrate D-Melezitose C-Source, carbohydrate MaltitolC-Source, carbohydrate a-Mehtyl-D-Galactoside C-Source, carbohydrateb-Methyl-D-Galactoside C-Source, carbohydrate 3-Methylglucose C-Source,carbohydrate b-Methyl-D-Glucuronic acid C-Source, carboxylic acida-Methyl-D-Mannoside C-Source, carbohydrate b-Methyl-D-XylosideC-Source, carbohydrate Palatinose C-Source, carbohydrate D-RaffinoseC-Source, carbohydrate Salicin C-Source, carbohydrate SedoheptulosanC-Source, carbohydrate L-Sorbose C-Source, carbohydrate StachyoseC-Source, carbohydrate D-Tagatose C-Source, carbohydrate TuranoseC-Source, carbohydrate Xylitol C-Source, carbohydrateN-Acetyl-D-Glucosaminitol C-Source, carbohydrate g-Amino-B-Butyric acidC-Source, carboxylic acid d-Amino Valeric acid C-Source, carboxylic acidButyric acid C-Source, carboxylic acid Capric acid C-Source, carboxylicacid Caproic acid C-Source, carboxylic acid Citraconic acid C-Source,carboxylic acid Citramalic acid C-Source, carboxylic acid D-GlucosamineC-Source, carbohydrate 2-Hydroxybenzoic acid C-Source, carboxylic acid4-Hydroxybenzoic acid C-Source, carboxylic acid b-Hydroxybutyric acidC-Source, carboxylic acid g-Hydroxybutyric acid C-Source, carboxylicacid a-Keto-Valeric acid C-Source, carboxylic acid Itaconic acidC-Source, carboxylic acid 5-Keto-D-Gluconic acid C-Source, carboxylicacid D-Lactic acid Methyl Ester C-Source, ester Malonic acid C-Source,carboxylic acid Melibionic acid C-Source, carbohydrate Oxalic acidC-Source, carboxylic acid Oxalomalic acid C-Source, carboxylic acidQuinic acid C-Source, carboxylic acid D-Ribono-1,4-Lactone C-Source,carboxylic acid Sebacic acid C-Source, carboxylic acid Sorbic acidC-Source, carboxylic acid Succinamic acid C-Source, carboxylic acidD-Tartaric acid C-Source, carboxylic acid L-Tartaric acid C-Source,carboxylic acid Acetamide C-Source, amide L-Alaninamide C-Source, amideN-Acetyl-L-Glutamic acid C-Source, amino acid L-Arginine C-Source, aminoacid Glycine C-Source, amino acid L-Histidine C-Source, amino acidL-Homoserine C-Source, amino acid Hydroxy-L-Proline C-Source, amino acidL-Isoleucine C-Source, amino acid L-Leucine C-Source, amino acidL-Lysine C-Source, amino acid L-Methionine C-Source, amino acidL-Ornithine C-Source, amino acid L-Phenylalanine C-Source, amino acidL-Pyroglutamic acid C-Source, amino acid L-Valine C-Source, amino acidD,L-Carnitine C-Source, carboxylic acid sec0Butylamine C-Source, amineD,L-Octopamine C-Source, amine Putrescine C-Source, amineDihydroxyacetone C-Source, alcohol 2,3-Butanediol C-Source, alcohol2,3-Butanedione C-Source, alcohol 3-Hydroxy-2-butanone C-Source, alcohol

TABLE 5 Bacterial Species Detected at Low Frequency in Vaginal Samplesfrom Vancomycin-Treated Mice Mean Median abundance abundance day day 6(out of 6 (out of Site Group Taxonomy 10,000) 10,000) vaginal VancomycinKF008552.1.1432 D_0__Bacteria; 0.291242675 0.024255713D_1__Proteobacteria; D_2__Gammaproteobacteria; D_3__Enterobacteriales;D_4__Enterobacteriaceae; D_5__Klebsiella; D_6__Klebsiella penumoniaevaginal Vancomycin AB740357.1.1462 D_0__Bacteria; 1.436524722 0D_1__Proteobacteria; D_2__Gammaproteobacteria; D_3__Enterobacteriales;D_4__Enterobacteriaceae; D_5__Pantoea; D_6__Pantoea sp. NCCP-532 vaginalVancomycin DQ799428.1.1372 D_0__Bacteria; 0.348310693 0D_1__Verrucomicrobia; D_2__Verrucomicrobiae; D_3__Verrucomicrobiales;D_4__Verrucomicrobiaceae; D_5__Akkermansia; D_6__uncultured bacteriumvaginal Vancomycin JX094996.1.1390 D_0__Bacteria; 0.349310693 0D_1__Firmictues; D_2__Clostridia; D_3__Clostridiales;D_4__Lachnospiraceae; D_5__Blautia; D_6_uncultured bacterium vaginalVancomycin EU459716.1.1286 D_0__Bacteria; 0.348310693 0 D_1__Firmicutes;D_2__Clostridia; D_3__Clostridiales; D_4__Lachnospiraceae;D_5__uncultured; D_6__uncultured bacterium vaginal VancomycinEU457230.1.1391 D_0__Bacteria; 0.696621386 0 D_1__Firmictues;D_2__Clostridia; D_3__Clostridiales; D_4__Lachnospiraceae; D_5__IncertaeSedis; D_6__uncultured bacterium vaginal Vancomycin EU459317.1.1373D_0__Bacteria; 0.348310693 0 D_1__Firmicutes; D_2__Clostridia;D_3__Clostridiales; D_4__Clostridiaceae 1; D_5__Clostridium sensuctricto 1; D_6__uncultured bacterium vaginal Vancomycin HM817954.1.1353D_0__Bacteria; 0.348310693 0 D_1__Firmicutes; D_2__Clostridia;D_3__Clostridiales; D_4__Lachnospiraceae; D_5__Roseburia;D_6__uncultured bacterium vaginal Vancomycin GQ134873.1.1373D_0__Bacteria; 0.348310693 0 D_1__Flirmicutesl D_2__Clostridia;D_3__Clostridiales; D_4__Clostridiaceae 1; D_5__Clostridium sensustricto 1; D_6__uncultured bacterium vaginal Vancomycin FJ879074.1.1494D_0__Bacteria; 0.348310693 0 D_1__Firmicutes; D_2__Clostridia;D_3__Clostridiales; D_4__Lachnospiraceae; D_5__uncultured;D_6__uncultured bacterium vaginal Vancomycin EU774816.1.1381D_0__Bacteria; 0.348310693 0 D_1__Firmicutes; D_2__Clostridia;D_3__Clostriadiales; D_4__Clostridiaceae 1; D_5__Clostridium sensustricto 1; D_6__uncultured bacterium vaginal Vancomycin EU775614.1.1398D_0__Bacteria; 0.417063419 0 D_1__Proteobacteria;D_2__Gammaproteobacteria; D_3__Enterobacteriales;D_4__Enterobacteriaceae; D_5__Enterobaacter; D_6__uncultured bacterium

Unless otherwise indicated, all numbers expressing quantities ofingredients, reaction conditions, and so forth used in thespecification, including claims, are to be understood as being modifiedin all instances by the term “about.” Accordingly, unless otherwiseindicated to the contrary, the numerical parameters are approximationsand may vary depending upon the desired properties sought to beobtained. At the very least, and not as an attempt to limit theapplication of the doctrine of equivalents to the scope of the claims,each numerical parameter should be construed in light of the number ofsignificant digits and ordinary rounding approaches.

Unless otherwise indicated, the term “at least” preceding a series ofelements is to be understood to refer to every element in the series.

While the invention has been particularly shown and described withreference to a preferred embodiment and various alternate embodiments,it will be understood by persons skilled in the relevant art thatvarious changes in form and details can be made therein withoutdeparting from the spirit and scope of the invention.

All references, issued patents and patent applications cited within thebody of the instant specification are hereby incorporated by referencein their entirety, for all purposes.

The foregoing description of the embodiments of the invention has beenpresented for the purpose of illustration; it is not intended to beexhaustive or to limit the invention to the precise forms disclosed.Persons skilled in the relevant art can appreciate that manymodifications and variations are possible in light of the abovedisclosure. Finally, the language used in the specification has beenprincipally selected for readability and instructional purposes, and itmay not have been selected to delineate or circumscribe the inventivesubject matter. It is therefore intended that the scope of the inventionbe limited not by this detailed description, but rather by any claimsthat issue on an application based hereon. Accordingly, the disclosureof the embodiments of the invention is intended to be illustrative, butnot limiting, of the scope of the invention, which is set forth in thefollowing claims.

What is claimed is:
 1. A method of treating autoimmune or inflammatorydisease in a subject in need thereof, comprising administering to thesubject a pharmaceutical composition comprising a bacterial populationof a single bacterial species, wherein the single bacterial species isBlautia hydrogenotrophica, such that inflammation in the subject isreduced.
 2. The method of claim 1, wherein administering thepharmaceutical composition to the subject reduces secretion of at leastone pro-inflammatory cytokine by immune cells of the subject.
 3. Themethod of claim 2, wherein the pro-inflammatory cytokine is selectedfrom the group consisting of IL-1a, IL-6, IFNγ and TNF alpha.
 4. Themethod of claim 2, wherein the immune cells are peripheral bloodmononuclear cells (PBMCs).
 5. The method of claim 1, whereinadministering the pharmaceutical composition to the subject increasessecretion of at least one anti-inflammatory cytokine by immune cells ofthe subject.
 6. The method of claim 5, wherein the anti-inflammatorycytokine is selected from the group consisting of IL-4, IL-5, IL-9,IL-10 and IL-13.
 7. The method of claim 5, wherein the immune cells areperipheral blood mononuclear cells (PBMCs).
 8. The method of claim 1,wherein administering the pharmaceutical composition to the subjectincreases the proportion of regulatory T cells in the subject.
 9. Themethod of claim 1, wherein the autoimmune or inflammatory disease isselected from the group consisting of inflammatory bowel disease (IBD),ulcerative colitis, Crohn's disease, multiple sclerosis (MS), systemiclupus erythematosus (SLE), type I diabetes, rheumatoid arthritis (RA),Sjögren's syndrome, uveitis, and Celiac disease.